Tibial Growth Plate Research Articles

Lactiplantibacillus plantarum P101 Ameliorates TiO2 NP-Induced Bone Injury in Young Rats by Remodeling the Gut Microbiota and Inhibiting the Production of Pro-Inflammatory Cytokines.

To evaluate the therapeutic effect of oral administration of Lactiplantibacillus plantarum P101 (P101) on skeletal injury in young rats exposed to titanium dioxide nanoparticles (TiO2 NPs), and explore the potential mechanism. Four-week-old male rats were orally administration to TiO2 NPs and supplemented with P101 2hours later for 4weeks. The growth and development, food intake, bone metabolism and serum inflammatory markers of the rats were evaluated. Their tibias were observed and evaluated using microcomputed tomography (micro-CT), tartrate-resistant acid phosphatase (TRAP) staining, immunohistochemistry (IHC) and real-time quantitative PCR (RT-qPCR). We observed the tibia growth plate using safranin and fast green staining. 16S rDNA sequence analysis of fecal samples was performed to observe changes in the gut microbiota. Our results showed that TiO2 NPs can lead to bone growth inhibition and osteoporosis, induce intestinal flora imbalance, and induce inflammation in young rats. Further mechanistic studies suggested that TiO2 NPs disrupts intestinal flora and increases serum IL-1β levels, which increased the expression of RANKL in bone, thereby enhancing osteoclast differentiation and function, leading to bone loss. Through a P101 supplementation experiment, we found that P101 ameliorated the inflammation and osteoporosis on bone caused by TiO2 NPs. This study showed that the mechanism by which P101 alleviates bone damage caused by TiO2 NPs may be through restoring intestinal microbial homeostasis and inhibiting inflammatory response.

CCR7 depletion alleviates bony growth imbalance following physeal injury in mice

Growth plates are the frequent sites of skeletal injury in children, leading to skeletal growth imbalances. Chemokines, including the receptor CCR7, play a crucial role in stem cell recruitment and cartilage homeostasis, with previous studies linking CCR7 to osteoarthritis progression. However, its role in growth plate cartilage remains unclear. We analyzed the role of CCR7 in the physeal cartilage repair process in mice model. Physeal injury was created in the proximal tibia in 3-week-old C57BL/6 mice (WT) and CCR7-knockout mice (CCR7−/−). Tibial length was measured macroscopically and sections of the physeal injury were analyzed histologically and immunohistochemically. Height and bone volume of the tibial growth plate and bone mineral density (BMD) of the subchondral area were measured by micro-CT. Mesenchymal stem cells (MSCs) were harvested and gene expression after osteogenic differentiation was analyzed using qRT-PCR. At 1, 3 and 5 weeks postoperatively, injured tibiae of CCR7−/− mice were less shortened than those of WT mice. Bone volume of the physeal bridge was significantly lower in CCR7−/− mice than in WT mice. In contrast, BMD of the subchondral area was comparable between CCR−/− and WT mice, and between sham and operated tibiae. In osteogenic differentiation, CCR7−/− mice showed significantly lowered expression of osteogenic markers such as Osterix, Runx2 and Type X collagen. We demonstrated CCR7 depletion in mice inhibited physeal bridge formation and ameliorated growth imbalances after physeal injury.

7999 Tumor Induced Osteomalacia Caused By Glomangiopericytoma

Abstract Disclosure: N. Kabir: None. E. Campbell: None. S. Singh: None. Introduction: Tumor induced osteomalacia (TIO) is a rare paraneoplastic syndrome caused primarily by FGF-23 secreting tumors. Excessive production of FGF23 causes hypophosphatemia, decreased phosphate resorption in the gut, increased renal phosphate wasting, and decreased activation of Vitamin D. This leads to decreased bone density, bone pain, proximal muscle weakness, impaired gait, and recurrent fractures. Case Presentation: 44 year old male with a history of recurrent insufficiency fractures and weakness. At the time of evaluation, he reported fracture of tibial growth plate, sacral fracture, and metatarsal stress fracture. Other symptoms included bone pain, fatigue, muscle weakness, and intercostal muscle spasms with exertion. His pain was debilitating, requiring crutches to ambulate. Available labs at that time were significant for hypophosphatemia (1.3 mg/dL), elevated ALP (160 U/L), low 25OH-vitamin D (26 ng/mL), and normal calcium (8.7 mg/dL). Urine studies demonstrated phosphaturia (fractional excretion of phosphate, 62.4%). FGF23 level resulted at the upper limit of normal (210 RU/mL). Pending further evaluation, burosumab was started. Further work up included a 68Ga-PET-DOTATATE scan revealing a somatostatin receptor expressing soft tissue mass in the left sphenoid sinus and posterior left ethmoid sinus. The scan also demonstrated impending bilateral femur fractures, prior rib fractures, and sacral insufficiency fractures. Based on these findings, he was admitted to the hospital and underwent prophylactic, intramedullary nail fixation of his bilateral femur fractures. He underwent resection of skull base tumor with the final pathology revealing a sinonasal glomangiopericytoma. Immunohistochemical stain was positive for CD99 and cyclin D1, and focally positive for SMA, consistent with histopathologic diagnosis. Patient required prolonged inpatient rehabilitation to recover independent ambulation. Discussion: Sinonasal glomangiopericytoma (SNGP) is a mesenchymal tumor that accounts for <0.5% of all nasal and sinus tumors. TIO due to SNGP is extremely rare, with only a few cases reported since the 1990s. As seen with our patient, TIO from SNGP rarely presents with sinonasal symptoms, and can often be overlooked. Our patient hadb significant progression of disease and disability due to numerous delays in diagnosis and management. Definitive cure for TIO can be achieved with complete surgical resection of the tumor, but failure to diagnose in a timely manner can result in decline in quality of life. Our takeaway from this case is that providers should obtain FGF23 levels, especially in the setting of hypophosphatemia and phosphaturia, to complete the work up for osteomalacia. Presentation: 6/1/2024

Adult Tillaux-Chaput tubercle fracture of the ankle with rupture of the peroneus tertius muscle: a case report

BackgroundTillaux-Chaput tubercle fractures occur in adolescents, which are often associated with the separation of the distal tibial growth plate. These types of fractures are rare in adults and even rarer when accompanied by a rupture of the peroneus tertius muscle. Given the limited number of reported cases, there is limited clinical awareness, resulting in missed diagnoses and delayed treatment, ultimately affecting ankle function.Case presentationWe report a case of an adult patient who experienced a right ankle injury resulting in swelling and pain after a traffic accident. Initial examination failed to identify the rupture of the peroneus tertius muscle, but the patient was observed to have restricted dorsiflexion and eversion of the foot. Surgical exploration through an anterolateral incision confirmed the rupture and the muscle was then repaired. The patient received four weeks of cast immobilization and then engaged in progressive rehabilitation exercises.Discussion and conclusionThis report shares the diagnostic and therapeutic experiences of an adult with a Tillaux-Chaput tubercle fracture associated with peroneus tertius muscle rupture to improve clinical recognition of such injuries, thus preventing misdiagnosis and treatment delays.

Tibial Damage Caused by T-2 Toxin in Goslings: Bone Dysplasia, Poor Bone Quality, Hindered Chondrocyte Differentiation, and Imbalanced Bone Metabolism.

T-2 toxin, the most toxic type A trichothecene, is widely present in grain and animal feed, causing growth retardation and tissue damage in poultry. Geese are more sensitive to T-2 toxin than chickens and ducks. Although T-2 toxin has been reported to cause tibial growth plate (TGP) chondrodysplasia in chickens, tibial damage caused by T-2 toxin in geese has not been fully demonstrated. This study aims to investigate the adverse effects of T-2 toxin on tibial bone development, bone quality, chondrocyte differentiation, and bone metabolism. Here, forty-eight one-day-old male Yangzhou goslings were randomly divided into four groups and daily gavaged with T-2 toxin at concentrations of 0, 0.5, 1.0, and 2.0 mg/kg body weight for 21 days, respectively. The development of gosling body weight and size was determined by weighing and taking body measurements after exposure to different concentrations of T-2 toxin. Changes in tibial development and bone characteristics were determined by radiographic examination, phenotypic measurements, and bone quality and composition analyses. Chondrocyte differentiation in TGP and bone metabolism was characterized by cell morphology, tissue gene-specific expression, and serum marker levels. Results showed that T-2 toxin treatment resulted in a lower weight, volume, length, middle width, and middle circumference of the tibia in a dose-dependent manner (p < 0.05). Moreover, decreased bone-breaking strength, bone mineral density, and contents of ash, Ca, and P in the tibia were observed in T-2 toxin-challenged goslings (p < 0.05). In addition, T-2 toxin not only reduced TGP height (p < 0.05) but also induced TGP chondrocytes to be disorganized with reduced numbers and indistinct borders. As expected, the apoptosis-related genes (CASP9 and CASP3) were significantly up-regulated in chondrocytes challenged by T-2 toxin with a dose dependence, while cell differentiation and maturation-related genes (BMP6, BMP7, SOX9, and RUNX2) were down-regulated (p < 0.05). Considering bone metabolism, T-2 toxin dose-dependently and significantly induced a decreased number of osteoblasts and an increased number of osteoclasts in the tibia, with inhibited patterns of osteogenesis-related genes and enzymes and increased patterns of osteoclast-related genes and enzymes (p < 0.05). Similarly, the serum Ca and P concentrations and parathyroid hormone, calcitonin, and 1, 25-dihydroxycholecalciferol levels decreased under T-2 toxin exposure (p < 0.05). In summary, 2.0 mg/kg T-2 toxin significantly inhibited tibia weight, length, width, and circumference, as well as decreased bone-breaking strength, density, and composition (ash, calcium, and phosphorus) in 21-day-old goslings compared to the control and lower dose groups. Chondrocyte differentiation in TGP was delayed by 2.0 mg/kg T-2 toxin owing to cell apoptosis. In addition, 2.0 mg/kg T-2 toxin promoted bone resorption and inhibited osteogenesis in cellular morphology, gene expression, and hormonal modulation patterns. Thus, T-2 toxin significantly inhibited tibial growth and development with a dose dependence, accompanied by decreased bone geometry parameters and properties, hindered chondrocyte differentiation, and imbalanced bone metabolism.

Intermittent mechanical loading on mouse tibia accelerates longitudinal bone growth by inducing PTHrP expression in the female tibial growth plate.

It is not clear as to whether weight bearing and ambulation may affect bone growth. Our goal was to study the role of mechanical loading (one of the components of ambulation) on endochondral ossification and longitudinal bone growth. Thus, we applied cyclical, biologically relevant strains for a prolonged time period (4 weeks) to one tibia of juvenile mice, while using the contralateral one as an internal control. By the end of the 4-week loading period, the mean tibial growth of the loaded tibiae was significantly greater than that of the unloaded tibiae. The mean height and the mean area of the loaded tibial growth plates were greater than those of the unloaded tibiae. In addition, in female mice we found a greater expression of PTHrP in the loaded tibial growth plates than in the unloaded ones. Lastly, microCT analysis revealed no difference between loaded and unloaded tibiae with respect to the fraction of bone volume relative to the total volume of the region of interest or the tibial trabecular bone volume. Thus, our findings suggest that intermittent compressive forces applied on tibiae at mild-moderate strain magnitude induce a significant and persistent longitudinal bone growth. PTHrP expressed in the growth plate appears to be one growth factor responsible for stimulating endochondral ossification and bone growth in female mice.

Case Report: Halting terminal osseous overgrowth post tibia amputation in children: a report of three cases

Terminal osseous overgrowth is a common complication after trans-diaphyseal amputation in children, leading to pain, soft tissue problems, and recurrent surgical procedures. We report three different cases with post-amputation issues of osseous overgrowth, ulceration, and deformity over the amputation site. The first case involves a 9-year-old boy with a right leg congenital amputation secondary to amniotic band syndrome. The right below-knee stump later experienced recurrent episodes of osseous overgrowth, leading to ulceration. After the prominent tibia was resected and capped with the ipsilateral proximal fibula, a positive outcome was achieved with no more recurrent overgrowth over the right leg stump. The second case involves a 9-year-old girl born with an amniotic constriction band over both legs. Her left leg remained functional after a circumferential Z-plasty, but the right leg was a congenital below-knee amputation. Multiple refashioning surgeries were performed on the right leg due to osseous overgrowth but the patient continued to experience recurrent overgrowth causing pain and difficulty fitting into a prosthesis. We performed osteocartilaginous transfer of the proximal part of the ipsilateral fibula to the right tibial end, successfully preventing the overgrowth of the tibia without any complications. The third case involves an 11-year-old boy with a history of meningococcal septicemia who underwent a right below-knee amputation and left ankle disarticulation due to complications of septic emboli. He experienced a prominent right distal tibia stump, which later developed into valgus deformity as a result of the previous insult to the proximal tibial growth plate. We performed a corrective osteotomy over the proximal right tibia and capped the entire tibia with the ipsilateral fibula as an intramedullary splint for the osteotomy site. Post-operatively, we achieved satisfactory deformity correction and successfully halted the recurrent overgrowth over the right tibia stump. The method of ipsilateral fibula capping is safe and effective in managing the osseous overgrowth complications in trans-diaphyseal amputations among children. Therefore, it is a reasonable option during primary below-knee amputations in children compared to multiple refashioning surgeries.

Age-related changes in densitometry and histomorphometry of long bone during the pubertal growth spurt in male rats.

Because experimental studies to determine the developmental toxicity of exposure to various substances in children are impossible, many studies use immature male rats. This study aimed to provide normative data for longitudinal bone growth with age during the puberty in male rats. In order to evaluate long bone growth and mineralization we examined bone size and bone density by dual-energy X-ray absorptiometry, analyzed histomorphometry of the growth plate, and serum hormone levels relevant to bone growth from postnatal day (PD)20 to PD60. The length and weight of long bones increased strongly by PD40, and no further increase was observed after PD50. On the other hand, tibial growth plate height decreased sharply after PD50 along with a reduction in the number of cells and columns, which was probably responsible for the absence of further lengthening of long bones. Parameters related to bone formation such as bone area ratio, and the thickness and number of trabeculae, also increased significantly between PD40 and PD50. Furthermore, serum levels of IGF-1 peaked at PD30 and testosterone increased rapidly on and after PD40, when IGF-1 levels were going down. These changes may participate in the parallel increase in mineral acquisition, as well as lengthening of long bones. Our findings provide comprehensive data for changes in bone density, histomorphometry of long bones, and hormone levels relevant to bone growth during the growth spurt. This will be useful for planning animal toxicological studies, particularly for deciding on the appropriate age of animals to use in given experiments.

Impact of the arginine silicate inositol complex on bone metabolism in broiler chickens with tibial dyschondroplasia caused by manganese deficiency

ABSTRACT 1. Tibial dyschondroplasia (TD) is a skeletal disorder in broilers that has financial implications, necessitating dietary modifications to reduce the prevalence of this disease. This study explored how arginine silicate inositol complex (ASI) supplementation affected tibial growth plate (TGP) and overall bone health in broilers with manganese (Mn) deficiency-induced TD. 2. A total of 240 broiler chicks were divided into four groups, each consisting of 60 birds (15 replicates of four broilers each) as follows: i) Control, with 60 mg Mn per kg of diet; ii) ASI, with 60 mg Mn and 1 g ASI per kg of diet; iii) TD, with 22 mg Mn per kg of diet, and iv) TD+ASI, with 22 mg Mn and 1 g ASI per kg of diet. 3. It was found that ASI supplementation increased tibial bone length in Mn-deficient TD broilers (p = 0.007). There was no Mn x ASI interaction for other bone morphometry variables (p > 0.05). However, both tibial bone mineral content and density were affected by Mn and ASI (p < 0.05). With ASI supplementation, serum bone-specific alkaline phosphatase and osteocalcin levels were elevated in the TD+ASI group compared to the TD group (p < 0.001). In the TD group, osteoprotegerin (OPG) levels in the TGP decreased compared to the control groups (p < 0.001). 4. In contrast, ASI supplementation in the TD broilers counteracted the decrease in OPG compared to TD broilers without ASI supplementation (p < 0.001). The Mn level and ASI supplementation significantly influenced the OPG/receptor activator of the nuclear factor-κB ligand ratio (p < 0.001). 5. In conclusion, the results demonstrated that inclusion of ASI in broiler diets could enhance bone formation variables by controlling OPG levels in the TGP, potentially serving as an effective method to decrease the occurrence of TD.

Subperiosteal delivery of transforming growth factor beta 1and human growth hormone from mineralized PCL films.

The ability to locally deliver bioactive molecules to distinct regions of the skeleton may provide a novel means by which to improve fracture healing, treat neoplasms or infections, or modulate growth. In this study, we constructed single-sided mineral-coated poly-ε-caprolactone membranes capable of binding and releasing transforming growth factor beta 1 (TGF-β1) and human growth hormone (hGH). After demonstrating biological activity in vitro and characterization of their release, these thin bioabsorbable membranes were surgically implanted using an immature rabbit model. Membranes were circumferentially wrapped under the periosteum, thus placed in direct contact with the proximal metaphysis to assess its bioactivity in vivo. The direct effects on the metaphyseal bone, bone marrow, and overlying periosteum were assessed using radiography and histology. Effects of membrane placement at the tibial growth plate were assessed via physeal heights, tibial growth rates (pulsed fluorochrome labeling), and tibial lengths. Subperiosteal placement of the mineralized membranes induced greater local chondrogenesis in the plain mineral and TGF-β1 samples than the hGH. More exuberant and circumferential ossification was seen in the TGF-β1 treated tibiae. The TGF-β1 membranes also induced hypocellularity of the bone marrow with characteristics of gelatinous degeneration not seen in the other groups. While the proximal tibial growth plates were taller in the hGH treated than TGF-β1, no differences in growth rates or overall tibial lengths were found. In conclusion, these data demonstrate the feasibility of using bioabsorbable mineral coated membranes to deliver biologically active compounds subperiosteally in a sustained fashion to affect cells at the insertion site, bone marrow, and even growth plate.

Skeletal abnormalities in mice with Dnmt3a missense mutations

Overgrowth and intellectual disability disorders in humans are typified by length/height and/or head circumference ≥ 2 standard deviations above the mean as well as intellectual disability and behavioral comorbidities, including autism and anxiety. Tatton-Brown-Rahman Syndrome is one type of overgrowth and intellectual disability disorder caused by heterozygous missense mutations in the DNA methyltransferase 3A (DNMT3A) gene. Numerous DNMT3A mutations have been identified in Tatton-Brown-Rahman Syndrome patients and may be associated with varying phenotype severities of clinical presentation. Two such mutations are the R882H and P904L mutations which result in severe and mild phenotypes, respectively. Mice with paralogous mutations (Dnmt3aP900L/+ and Dnmt3aR878H/+) exhibit overgrowth in their long bones (e.g., femur, humerus), but the mechanisms responsible for their skeletal overgrowth remain unknown. The goal of this study is to characterize skeletal phenotypes in mouse models of Tatton-Brown-Rahman Syndrome and identify potential cellular mechanisms involved in the skeletal overgrowth phenotype. We report that mature mice with the Dnmt3aP900L/+ or Dnmt3aR878H/+ mutation exhibit tibial overgrowth, cortical bone thinning, and weakened bone mechanical properties. Dnmt3aR878H/+ mutants also contain larger bone marrow adipocytes while Dnmt3aP900L/+ mutants show no adipocyte phenotype compared to control animals. To understand the potential cellular mechanisms regulating these phenotypes, growth plate chondrocytes, osteoblasts, and osteoclasts were assessed in juvenile mutant mice using quantitative static histomorphometry and dynamic histomorphometry. Tibial growth plates appeared thicker in mutant juvenile mice, but no changes were observed in osteoblast activity or osteoclast number in the femoral mid-diaphysis. These studies reveal new skeletal phenotypes associated with Tatton-Brown-Rahman Syndrome in mice and provide a rationale to extend clinical assessments of patients with this condition to include bone density and quality testing. These findings may be also informative for skeletal characterization of other mouse models presenting with overgrowth and intellectual disability phenotypes.

Microwave Ablation of the Pig Growth Plate: Proof of Concept for Minimally Invasive Epiphysiodesis.

Different surgical methods for epiphysiodesis of limb length discrepancy (LLD) have been described. Although these methods are variably effective, they are associated with morbidity (pain and limp) and potential complications. Microwave ablation is a less-invasive opportunity to halt growth by selectively destroying the growth plate via thermal energy to treat LLD in children. In this proof-of-concept study using an in vivo pig model, we asked: (1) What is the durability of response 2 to 4 months after microwave ablation of the tibial growth plate as measured by length and angulation of the tibia via a CT scan? (2) Was articular cartilage maintained as measured by standard histologic staining for articular cartilage viability? To develop an in vivo protocol for microwave ablation, we placed microwave antennas adjacent to the proximal tibia growth plate in the cadaveric hindlimbs of 18 3-month-old pigs. To determine the suitable time, we varied ablation from 90 to 270 seconds at 65-W power settings. After sectioning the tibia, we visually assessed for discoloration (implying growth plate destruction) that included the central growth plate but did not encroach into the epiphysis in a manner that could disrupt the articular surface. Using this information, we then performed microwave ablation on three live female pigs (3.5 to 4 months old) to evaluate physiologic changes and durability of response. A postprocedure MRI was performed to ensure the intervention led to spatial growth plate alterations similar to that seen in cadavers. This was followed by serial CT, which was used to assess the potential effect on local bone and growth until the animals were euthanized 2 to 4 months after the procedure. We analyzed LLD, angular deformity, and bony deformity using CT scans of both tibias. The visibility of articular cartilage was compared with that of the contralateral tibia via standard histologic staining, and growth rates of the proximal tibial growth plate were compared via fluorochrome labeling. Eighteen cadaveric specimens showed ablation zones across the growth plate without visual damage to the articular surface. The three live pigs did not exhibit changes in gait or require notable pain medication after the procedure. Each animal demonstrated growth plate destruction, expected limb shortening (0.8, 1.2, and 1.5 cm), and bony cavitation around the growth plate. Slight valgus bone angulation (4º, 5º, and 12º) compared with the control tibia was noted. No qualitatively observable articular cartilage damage was encountered from the histologic comparison with the contralateral tibia for articular cartilage thickness and cellular morphology. A microwave antenna placed into a pig's proximal tibia growth plate can slow the growth of the tibia without apparent pain and alteration of gait and function. Further investigation and refinement of our animal model is ongoing and includes shorter ablation times and comparison of dynamic ablation (moving the antennae during the ablation) as well as static ablation of the tibia from a medial and lateral portal. These refinements and planned comparison with standard mechanical growth arrest in our pig model may lead to a similar approach to ablate growth plates in children with LLD.

Longitudinal skeletal growth and growth plate morphological characteristics of chondro-tissue specific CUL7 knockout mice

Background3 M syndrome is first reported in 1975，which characterized by severe pre- and postnatal growth retardation, skeletal malformation and facial dysmorphism. These three genes (CUL7, OBSL1 and CCDC8) have been identified to be respond for 3 M syndrome, of which CUL7 is accounting for approximately 70%. To date, the molecular mechanism underlying the pathogenesis of 3 M syndrome remains poorly understood. Previous studies showed that no Cul7-/- mice could survive after birth, because of growth retardation at late gestational stage and respiratory distress after birth. The establishment of the animal model of cartilage specific Cul7 knockout mice (Cul7fl/fl;Col2a1-CreERT2 mice) has confirmed that Cul7fl/fl;Col2a1-CreERT2 mice can be selective in a time- and tissue-dependent manner, which can provide an experimental basis for further research on severe genetic diseases related to growth plates. ObjectiveTo establish a model of Cul7fl/fl;Col2a1-CreERT2 mice based on Cre/LoxP system, and to further observe its phenotype and morphological changes in growth plate. MethodsThe Cul7fl/fl;Col2a1-CreERT2 mice were taken as the experimental group, while the genotype of Cul7fl/+;Col2a1-CreERT2 mice were used as the control group. The gross morphological features and X-ray films of limbs in the two groups were observed every week for 3–6 consecutive weeks, and the length of the mice from nose to the tail, the length of femur and tibia were recorded. In the meantime, The histological morphology of tibial growth plates was compared between the two groups. ResultsA preliminary model of Cul7fl/fl;Col2a1-CreERT2 mice was established. The Cul7fl/fl;Col2a1-CreERT2 mice had abnormally short and deformed limbs (P<0.05), increased thickness of growth plate, the disorderly arranged chondrocyte columns, decreased number of cells in the proliferation zone, changes in the shape from flat to round, obviously expanded extracellular matrix, and disordered arrangement, thickening and loosening of bone trabecula at the proximal metaphysis of the femur. ConclusionsThe knockout of Cul7 gene may affect both the proliferation of chondrocytes and the endochondral osteogenesis, confirming that Cul7 is essential for the normal development of bone in the body.

Adaptations of bone and bone vasculature to muscular stretch training.

The magnitude of bone formation and remodeling is linked to both the magnitude of strain placed on the bone and the perfusion of bone. It was previously reported that an increase in bone perfusion and bone density occurs in the femur of old rats with moderate aerobic exercise training. This study determined the acute and chronic effects of static muscle stretching on bone blood flow and remodeling. Old male Fischer 344 rats were randomized to either a naive or stretch-trained group. Static stretching of ankle flexor muscles was achieved by placement of a dorsiflexion splint on the left ankle for 30min/d, 5d/wk for 4wk. The opposite hindlimb served as a contralateral control (nonstretched) limb. Bone blood flow was assessed during and after acute stretching in naive rats, and at rest and during exercise in stretch-trained rats. Vascular reactivity of the nutrient artery of the proximal tibia was also assessed in stretch-trained rats. MicroCT analysis was used to assess bone volume and micro-architecture of the trabecular bone of both tibias near that growth plate. In naive rats, static stretching increased blood flow to the proximal tibial metaphasis. Blood flow to the proximal tibial metaphysis during treadmill exercise was higher in the stretched limb after 4wk of daily stretching. Daily stretching also increased tibial bone weight and increased total volume in both the proximal and distal tibial metaphyses. In the trabecular bone immediately below the proximal tibial growth plate, total volume and bone volume increased, but bone volume/total volume was unchanged and trabecular connectivity decreased. In contrast, intravascular volume increased in this region of the bone. These data suggest that blood flow to the tibia increases during bouts of static stretching of the hindlimb muscles, and that 4wk of daily muscle stretching leads to bone remodeling and an increase in intravascular volume of the tibial bone.

Targeting FGFR3 signaling and drug repurposing for the treatment of SLC26A2-related chondrodysplasia in mouse model

BackgroundMutations in Slc26a2 cause a spectrum of autosomal-recessive chondrodysplasia with a significant and negligible influence on the quality of life. It has been reported that Slc26a2 deficiency triggers the ATF6 branch of the UPR, which may, in turn, activate the negative regulator of the FGFR3 signaling pathway. However, the correlation between the deletion of Slc26a2 and the augmentation of downstream phosphorylation of FGFR3 has not been investigated in vivo. MethodsFirst, we constructed Slc26a2 and Fgfr3 double knockout mouse lines and observed gross views of the born mice and histological staining of the tibial growth plates. The second approach was to construct tamoxifen-inducible Cre-ERT2 mouse models to replicate SLC26A2-related non-lethal dysplastic conditions. Pharmacological intervention was performed by administering the FGFR3 inhibitor NVP-BGJ398. The effect of NVP-BGJ398 on chondrocytes was assessed by Alcian blue staining, proliferation, apoptosis, and chondrocyte-specific markers and then verified by western blotting for variations in the downstream markers of FGFR3. The growth process was detected using X-rays, micro-CT examination, histomorphometry staining of growth plates, and immunofluorescence. ResultsGenetic ablation of Fgfr3 in embryonic Slc26a2-deficient chondrocytes slightly attenuated chondrodysplasia. Subsequently, in the constructed mild dysplasia model, we found that postnatal intervention with Fgfr3 gene in Slc26a2-deficient chondrocytes partially alleviated chondrodysplasia. In chondrocyte assays, NVP-BGJ398 suppressed the defective phenotype of Slc26a2-deficient chondrocytes and restored the phosphorylation downstream of FGFR3 in a concentration-dependent manner. In addition, in vivo experiments showed significant alleviation of impaired chondrocyte differentiation, and micro-CT analysis showed a clear improvement in trabecular bone microarchitectural parameters. ConclusionOur results suggested that inhibition of FGFR3 signaling pathway overactivation and NVP-BGJ398 has promising therapeutic implications for the development of SLC26A2-related skeletal diseases in humans. The translational potential of this articleOur data provide genetic and pharmacological evidence that targeting FGFR3 signaling via NVP-BGJ398 could be a route for the treatment of SLC26A2-associated skeletal disorders, which promisingly advances translational applications and therapeutic development.

Altered Sox9 and FGF signaling gene expression in Aga2 OI mice negatively affects linear growth.

Osteogenesis imperfecta (OI), or brittle bone disease, is a disorder characterized by bone fragility and increased fracture incidence. All forms of OI also feature short stature, implying an effect on endochondral ossification. Using the Aga2+/- mouse, which has a mutation in type I collagen, we show an affected growth plate primarily due to a shortened proliferative zone. We used single-cell RNA-Seq analysis of tibial and femoral growth plate tissues to understand transcriptional consequences on growth plate cell types. We show that perichondrial cells, which express abundant type I procollagen, and growth plate chondrocytes, which were found to express low amounts of type I procollagen, had ER stress and dysregulation of the same unfolded protein response pathway as previously demonstrated in osteoblasts. Aga2+/- proliferating chondrocytes showed increased FGF and MAPK signaling, findings consistent with accelerated differentiation. There was also increased Sox9 expression throughout the growth plate, which is expected to accelerate early chondrocyte differentiation but reduce late hypertrophic differentiation. These data reveal that mutant type I collagen expression in OI has an impact on the cartilage growth plate. These effects on endochondral ossification indicate that OI is a biologically complex phenotype going beyond its known impacts on bone to negatively affect linear growth.

Substantially Delayed Maturation of Growth Plate Chondrocytes in "Humanized" PTH1R Mice with the H223R Mutation of Jansen's Disease.

Activating parathyroid hormone (PTH)/PTH-related Peptide (PTHrP) receptor (PTH1R) mutations causes Jansen's metaphyseal chondrodysplasia (JMC), a rare disease characterized by growth plate abnormalities, short stature, and PTH-independent hypercalcemia. Previously generated transgenic JMC mouse models, in which the human PTH1R allele with the H223R mutation (H223R-PTH1R) is expressed in osteoblasts via type Ia1 collagen or DMP1 promoters cause excess bone mass, while expression of the mutant allele via the type IIa1 collagen promoter results in only minor growth plate changes. Thus, neither transgenic JMC model adequately recapitulates the human disease. We therefore generated "humanized" JMC mice in which the H223R-PTH1R allele was expressed via the endogenous mouse Pth1r promoter and, thus, in all relevant target tissues. Founders with the H223R allele typically died within 2 months without reproducing; several mosaic male founders, however, lived longer and produced F1 H223R-PTH1R offspring, which were small and exhibited marked growth plate abnormalities. Serum calcium and phosphate levels of the mutant mice were not different from wild-type littermates, but serum PTH and P1NP were reduced significantly, while CTX-1 and CTX-2 were slightly increased. Histological and RNAscope analyses of the mutant tibial growth plates revealed markedly expanded zones of type II collagen-positive, proliferating/prehypertrophic chondrocytes, abundant apoptotic cells in the growth plate center and a progressive reduction of type X collagen-positive hypertrophic chondrocytes and primary spongiosa. The "humanized" H223R-PTH1R mice are likely to provide a more suitable model for defining the JMC phenotype and for assessing potential treatment options for this debilitating disease of skeletal development and mineral ion homeostasis. © 2023 The Authors. JBMR Plus published by Wiley Periodicals LLC on behalf of American Society for Bone and Mineral Research.

Is the ratio of the index to ring finger length, a potential marker for prenatal testosterone-estrogen balance, related to posterior tibial slope?

BackgroundThe 2D:4D ratio, an indicator of prenatal testosterone-estrogen balance, is the index finger (second finger) size's ratio to the ring finger (fourth finger) size. Asymmetric growth in the proximal tibial growth plate is considered to cause the increased posterior tibial slope (PTS) formation. Factors determining the amount of this asymmetry still need clarification. This study aims to evaluate the relationship between the 2D:4D ratio, which indicates the prenatal testosterone-estrogen balance, and the PTS. MethodMeeting the study criteria, 267 patients were included in the study. We measured the medial PTS (MPTS) and lateral PTS (LPTS) on MRI images and the lengths of the second and fourth fingers on both hands of the patients. Additionally, we compared 2D:4D ratios and MPTS and LPTS measurements. ResultsWe found a significant negative correlation between MPTS and the 2D:4D ratio, as well as between LPTS and the 2D:4D ratio in both hands (MPTS left vs. right hand: P < 0.001, r = −0.627 vs. P < 0.001, r = −0.498) (LPTS left vs. right hand: P < 0.001, r = 0.589 vs. P < 0.001, r = 0.404). Separately among males and females, there was a significant negative correlation between MPTS and the 2D:4D ratio, as well as between LPTS and the 2D:4D ratio in both hands (for males: MPTS left vs. right hand: P < 0.001, r = −0.607 vs. P < 0.001, r = −0.540)(for males: LPTS left vs.right hand: P < 0.001, r = 0.451 vs. P < 0.001, r = 0.406) (for females:MPTS left vs. right hand: P < 0.001, r = −0.638 vs. P < 0.001, r = −0.446) (for females:LPTS left vs.right hand: P < 0.001, r = 0.618 vs. P < 0.001, r = 0.403). ConclusionsThe 2D:4D ratio, an indicator of intrauterine testosterone-estrogen balance, is related to PTS. The effect of testosterone on the growth plate of the proximal tibia may be one of the factors determining the PTS value.Level of evidence: III retrospective comparative study.

Poster 246: Epidemiology of Tibial Spine Fractures in Children: A National Multicenter Study

Objectives: Tibial spine fractures (TSFs) are avulsion fractures at the insertion of the anterior cruciate ligament at the tibial intercondylar eminence. TSFs are rare injuries thought to occur most frequently in skeletally immature children aged 8 to 14 years. Historically, it was thought that the most common mechanism was falling off of bicycles, but recent research suggests a variety of additional mechanisms. Treatment for TSFs can be operative or nonoperative depending on fracture type/severity, concomitant injuries, and surgeon and patient/family preference. Estimates of the epidemiology of this patient population are limited to primarily single-center reports. The objective of this investigation was to characterize epidemiology, including demographic information, injury mechanism, fracture type, and selection of surgical or nonsurgical treatment, in this injury population across eight geographically distinct institutions using retrospective data. Methods: IRB and reliance approval was obtained for each participating hospital. Patients younger than 25 years treated for TSFs between January 1, 2000 and January 31, 2019 were included. Patients with multiple lower extremity fractures or other traumatic injuries were excluded. Data was collected regarding demographics, mechanisms of injury, imaging reports, treatment, and concomitant injuries. Data were presented as medians and interquartile ranges. Categorical data was presented as counts and percentages and was stratified by sex and/or Meyers and McKeever fracture type. Results: 661 patients met inclusion criteria. Median age at presentation was 12.2 years (IQR 9.7-14.1), with a higher prevalence in boys than girls (66.1%, 437/661). Diagnosis at a study center occurred at a median of 4.0 days (IQR 1.0-9.0 days) after injury. Of these patients, 71.6% (432/603) had first been seen elsewhere, most commonly by emergency department physicians (40.7%, 176/432). The most commonly reported injury mechanisms were contact (44.4%, 244/549) and twisting, non-contact (42.8%, 235/549), with hyperextension injuries representing a much smaller proportion at 3.6%. Injuries occurred most frequently during sports at 56.8% (355/625), which were primarily skiing (127/352, 36.1%) and football (97/352, 27.6%). Distal femur and proximal tibia growth plate status were classified as open in 67.1% and 72.6% of cases, respectively. Both growth plates were classified as closed in only 5.8% of cases. Meyers and McKeever fracture type was most commonly Type II (40.4%) or Type III (44.4%). Treatment was surgical in 83.3% (549/659) of cases, and of these, most were treated arthroscopically (ARIF) (470/549, 86.1%). Conclusions: This study represents, to our knowledge, the largest epidemiological report to date of pediatric patients with tibial spine fractures. TSFs were more common in males than females (66.1% male), and the median age at injury was 12.2 years, which falls within the commonly reported range of 8-14 years. Furthermore, only 5.8% of patients had closed regional physes, corroborating the theory that TSFs are more likely to occur in skeletally immature individuals. We believe the data presented in this study can be useful towards patient counselling by providing a comprehensive view of the common presentations of and treatment pathways for tibial spine fractures in the pediatric population. [Table: see text] [Table: see text][Table: see text]

The Effect of miR-140-5p with HDAC4 towards Growth and Differentiation Signaling of Chondrocytes in Thiram-Induced Tibial Dyschondroplasia.

There is evidence to suggest that microRNA-140-5p (miR-140), which acts as a suppressor, is often elevated and has a role in various malignancies. Nevertheless, neither the function nor the mechanisms in chondrocytes linked with bone disorders, e.g., tibial dyschondroplasia (TD), have been satisfactorily established. The purpose of this study was to look into the role of microRNA-140-5p (miR-140) and its interaction with HDAC4 in chondrocytes, as well as the implications for tibial dyschondroplasia (TD), with a particular focus on the relationship between low miR-140 expression and poor pathologic characteristics, as well as its physiological effects on chondrocyte growth, differentiation, and chondrodysplasia. In this investigation, we discovered that TD had a reduced expression level of the miR-140. There was a correlation between low miR-140 expression, poor pathologic characteristics, and the short overall survival of chondrocytes. Our findings show an aberrant reduction in miR-140 expression, and HDAC4 overexpression caused disengagement in resting and proliferation zones. This further resulted in uncontrolled cell proliferation, differentiation, and chondrodysplasia. Mechanistically, HDAC4 inhibited the downstream transcription factors MEF2C and Runx2 and interacted with Col-Ⅱ, Col-X, and COMP. However, miR-140 binding to the 3'-UTR of HDAC4 resulted in the growth and differentiation of chondrocytes. Moreover, the expression of HDAC4 through LMK-235 was significantly decreased, and the expression was significantly increased under ITSA-1, referring to a positive feedback circuit of miR-140 and HDAC4 for endochondral bone ossification. Furthermore, as a prospective treatment, the flavonoids of Rhizoma drynariae (TFRD) therapy increased the expression of miR-140. Compared to the TD group, TFRD treatment increased the expression of growth-promoting and chondrocyte differentiation markers, implying that TFRD can promote chondrocyte proliferation and differentiation in the tibial growth plate. Hence, directing this circuit may represent a promising target for chondrocyte-related bone disorders and all associated pathological bone conditions.