Novel pathogenic variants in SPARC as cause of osteogenesis imperfecta: Two case reports.

Pathogenic variants in SPARC cause a rare autosomal recessive form of osteogenesis imperfecta (OI), classified as OI type XVII, which was first reported in 2015. Only six patient cases with this specific form of OI have been reported to date. The SPARC protein plays a crucial role in the calcification of collagen in bone, synthesis of the extracellular matrix, and the regulation of cell shape. In this case report, we describe the phenotype of two patients with SPARC-related OI, including a patient with two novel pathogenic variants in the SPARC gene. Targeted Next Generation Sequencing revealed new compound heterozygous variants (c.484G > A p.(Glu162Lys)) and c.496C > T p.(Arg166Cys)) in one patient and a homozygous nonsense pathogenic variant (c.145C > T p.(Gln49*)) in the other. In line with previously reported cases, the two OI patients presented delayed motor development, muscular weakness, scoliosis, and multiple fractures. Interestingly, our study reports for the first time the occurrence of dentinogenesis imperfecta. The study also reports the effectiveness of bisphosphonate treatment for OI type XVII. This article enhances the genetic, clinical, therapeutic, and radiological understanding of SPARC-related OI.

Proteomic Analysis of Dupuytren's Contracture-Derived Sweat Glands Revealed the Synthesis of Connective Tissue Growth Factor and Initiation of Epithelial-Mesenchymal Transition as Major Pathogenetic Events.

Dupuytren's contracture (DC) is a chronic and progressive fibroproliferative disorder restricted to the palmar fascia of the hands. Previously, we discovered the presence of high levels of connective tissue growth factor in sweat glands in the vicinity of DC nodules and hypothesized that sweat glands have an important role in the formation of DC lesions. Here, we shed light on the role of sweat glands in the DC pathogenesis by proteomic analysis and immunofluorescence microscopy. We demonstrated that a fraction of sweat gland epithelium underwent epithelial-mesenchymal transition illustrated by negative regulation of E-cadherin. We hypothesized that the increase in connective tissue growth factor expression in DC sweat glands has both autocrine and paracrine effects in sustaining the DC formation and inducing pathological changes in DC-associated sweat glands.

Phenotypic Variation in Vietnamese Osteogenesis Imperfecta Patients Sharing a Recessive P3H1 Pathogenic Variant.

Osteogenesis imperfecta (OI) is a syndromic disorder of bone fragility with high variation in its clinical presentation. Equally variable is molecular aetiology; recessive forms are caused by approximately 20 different genes, many of which are directly implicated in collagen type I biosynthesis. Biallelic variants in prolyl 3-hydroxylase 1 (P3H1) are known to cause severe OI by affecting the competence of the prolyl 3-hydroxylation­cartilage associated protein­peptidyl-prolyl cis-trans isomerase B (P3H1-CRTAP-CyPB) complex, which acts on the Pro986 residue of collagen type I α 1 (COL1A1) and Pro707 collagen type I α 2 (COL1A2) chains. The investigation of an OI cohort of 146 patients in Vietnam identified 14 families with P3H1 variants. The c.1170+5G>C variant was found to be very prevalent (12/14) and accounted for 10.3% of the Vietnamese OI cohort. New P3H1 variants were also identified in this population. Interestingly, the c.1170+5G>C variants were found in families with the severe clinical Sillence types 2 and 3 but also the milder types 1 and 4. This is the first time that OI type 1 is reported in patients with P3H1 variants expanding the clinical spectrum. Patients with a homozygous c.1170+5G>C variant shared severe progressively deforming OI type 3: bowed long bones, deformities of ribcage, long phalanges and hands, bluish sclera, brachycephaly, and early intrauterine fractures. Although it remains unclear if the c.1170+5G>C variant constitutes a founder mutation in the Vietnamese population, its prevalence makes it valuable for the molecular diagnosis of OI in patients of the Kinh ethnicity. Our study provides insight into the clinical and genetic variation of P3H1-related OI in the Vietnamese population.

Personal protection equipment for orthopaedic and trauma surgery during the COVID-19 pandemic: The results of an EFORT survey initiative.

Orthopaedic and trauma surgeons performing surgery in the COVID-19 pandemic environment faced problems with availability, use, rationing, modification, compliance and recycling of personal protection equipment (PPE). Orthopaedic and trauma surgeons were not well informed concerning the use of PPE for aerosol-generating orthopaedic and trauma procedures. Scientific bodies, health authorities and management have provided insufficient guidelines for the use of PPE in aerosol-generating orthopaedic and trauma procedures. The availability of specific PPE for orthopaedic and trauma operating theatres is low. Hospital management and surgeons failed to address the quality of operating theatre ventilation or to conform to recommendations and guidelines. Operating theatre PPE negatively affected surgical performance by means of impaired vision, impaired communication, discomfort and fatigue. Existing PPE is not adequately designed for orthopaedic and trauma surgery, and therefore, novel or modified and improved devices are needed.

From late fatherhood to prenatal screening of monogenic disorders: evidence and ethical concerns.

BACKGROUND: With the help of ART, an advanced parental age is not considered to be a serious obstacle for reproduction anymore. However, significant health risks for future offspring hide behind the success of reproductive medicine for the treatment of reduced fertility associated with late parenthood. Although an advanced maternal age is a well-known risk factor for poor reproductive outcomes, understanding the impact of an advanced paternal age on offspring is yet to be elucidated. De novo monogenic disorders (MDs) are highly associated with late fatherhood. MDs are one of the major sources of paediatric morbidity and mortality, causing significant socioeconomic and psychological burdens to society. Although individually rare, the combined prevalence of these disorders is as high as that of chromosomal aneuploidies, indicating the increasing need for prenatal screening. With the help of advanced reproductive technologies, families with late paternity have the option of non-invasive prenatal testing (NIPT) for multiple MDs (MD-NIPT), which has a sensitivity and specificity of almost 100%. OBJECTIVE AND RATIONALE: The main aims of the current review were to examine the effect of late paternity on the origin and nature of MDs, to highlight the role of NIPT for the detection of a variety of paternal age-associated MDs, to describe clinical experiences and to reflect on the ethical concerns surrounding the topic of late paternity and MD-NIPT. SEARCH METHODS: An extensive search of peer-reviewed publications (1980-2021) in English from the PubMed and Google Scholar databases was based on key words in different combinations: late paternity, paternal age, spermatogenesis, selfish spermatogonial selection, paternal age effect, de novo mutations (DNMs), MDs, NIPT, ethics of late fatherhood, prenatal testing and paternal rights. OUTCOMES: An advanced paternal age provokes the accumulation of DNMs, which arise in continuously dividing germline cells. A subset of DNMs, owing to their effect on the rat sarcoma virus protein-mitogen-activated protein kinase signalling pathway, becomes beneficial for spermatogonia, causing selfish spermatogonial selection and outgrowth, and in some rare cases may lead to spermatocytic seminoma later in life. In the offspring, these selfish DNMs cause paternal age effect (PAE) disorders with a severe and even life-threatening phenotype. The increasing tendency for late paternity and the subsequent high risk of PAE disorders indicate an increased need for a safe and reliable detection procedure, such as MD-NIPT. The MD-NIPT approach has the capacity to provide safe screening for pregnancies at risk of PAE disorders and MDs, which constitute up to 20% of all pregnancies. The primary risks include pregnancies with a paternal age over 40 years, a previous history of an affected pregnancy/child, and/or congenital anomalies detected by routine ultrasonography. The implementation of NIPT-based screening would support the early diagnosis and management needed in cases of affected pregnancy. However, the benefits of MD-NIPT need to be balanced with the ethical challenges associated with the introduction of such an approach into routine clinical practice, namely concerns regarding reproductive autonomy, informed consent, potential disability discrimination, paternal rights and PAE-associated issues, equity and justice in accessing services, and counselling. WIDER IMPLICATIONS: Considering the increasing parental age and risks of MDs, combined NIPT for chromosomal aneuploidies and microdeletion syndromes as well as tests for MDs might become a part of routine pregnancy management in the near future. Moreover, the ethical challenges associated with the introduction of MD-NIPT into routine clinical practice need to be carefully evaluated. Furthermore, more focus and attention should be directed towards the ethics of late paternity, paternal rights and paternal genetic guilt associated with pregnancies affected with PAE MDs.

Serum sclerostin concentration is associated with specific adipose, muscle and bone tissue markers in lean adolescent females with increased physical activity.

OBJECTIVES: Sclerostin is an important regulator of bone mass involving the Wnt/ß-catenin signalling pathway. Relatively few studies have investigated the relationships of circulating sclerostin levels with adiposity-related and muscle-related biochemical factors in individuals with increased energy metabolism. The aim of this study was to investigate the associations of circulating sclerostin with adipokines, myokines, osteokines and body composition values in lean adolescent females with increased physical activity. METHODS: A total of 73 adolescent females who were physically active and aged 14-18 years old participated in the study. Sclerostin, leptin, resistin, tumour necrosis factor (TNF)-α, interleukin (IL)-6, irisin, osteocalcin, C-terminal telopeptide of type I collagen (CTx), insulin-like growth factor (IGF)-1 and insulin were obtained from fasting blood samples. Body composition was measured by dual-energy X-ray absorptiometry (DXA) and analyzed for body fat mass, lean body mass, bone mineral content and muscle mass. RESULTS: Serum sclerostin (117.9 ± 60.3 pg/mL) was correlated with age, age at menarche, body fat, muscle mass, training activity, leptin, TNF-α, irisin, osteocalcin, CTx and IGF-1. Multivariate linear regression analysis demonstrated that fat mass (ß = 0.434; p = 0.001), leptin (ß = -0.308; p = 0.015), irisin (ß = 0.227; p = 0.024) and CTx (ß = 0.290; p = 0.031) were the most important predictors of serum sclerostin concentration. CONCLUSIONS: Bone-derived sclerostin is associated with specific adipokine, myokine and osteokine values in lean adolescent females with increased physical activity. These results suggest that the interactions between bone, adipose and muscle tissues could also be associated with circulating sclerostin concentrations.

Sclerostin, preadipocyte factor-1 and bone mineral values in eumenorrheic adolescent athletes with different training patterns.

INTRODUCTION: The specific aims of the study were to compare possible differences in sclerostin and preadipocyte factor-1 (Pref-1) between rhythmic gymnasts (RG), swimmers (SW) and untrained controls (UC), and to investigate the relationships of sclerostin and Pref-1 with bone mineral characteristics in studied groups. MATERIALS AND METHODS: This study included 62 eumenorrheic adolescents (RG = 22; SW = 20; UC = 20). Bone mineral and body composition characteristics were measured by dual-energy X-ray absorptiometry, and sclerostin, Pref-1, osteocalcin and C-terminal telopeptide of type I collagen (CTx) were measured. RESULTS: Sclerostin was higher (P = 0.001) in RG (129.35 ± 51.01 pg/ml; by 74%) and SW (118.05 ± 40.05 pg/ml; by 59%) in comparison with UC (74.32 ± 45.41 pg/ml), while no differences (P = 0.896) were seen in Pref-1 (RG: 1.42 ± 0.16 ng/ml; SW: 1.41 ± 0.20 ng/ml; UC: 1.39 ± 0.26 ng/ml) between groups. Osteocalcin (RG: 7.74 ± 4.09 ng/ml; SW: 8.05 ± 4.18 ng/ml; UC: 7.04 ± 3.92 ng/ml; P = 0.843) and CTx (RG: 0.73 ± 0.22 ng/ml; SW: 0.64 ± 0.16 ng/ml; UC: 0.62 ± 0.20 ng/ml; P = 0.173) were not different between groups. Sclerostin correlated (P < 0.05) with whole-body bone mineral content (r = 0.61) and lumbar spine (LS) areal bone mineral density (aBMD) (r = 0.43) in RG, and femoral neck aBMD (r = 0.45) in UC. No correlation was found between sclerostin and bone mineral values in SW, and Pref-1 was not correlated with any bone mineral characteristics in studied groups. Sclerostin was the independent variable that explained 14% of the total variance (R2 × 100) in LS aBMD value only in RG. CONCLUSIONS: Adolescent athletes have higher sclerostin compared to UC. Sclerostin was correlated with bone mineral values and predicted areal bone mineral density in RG.

Alternative splicing of leptin receptor overlapping transcript in osteosarcoma.

IMPACT STATEMENT: Osteosarcoma (OS, also known as osteogenic sarcoma) is the most common primary malignancy of bone in children and adolescents. The molecular mechanisms of OS are extremely complicated and its molecular mediators remain to be elucidated. We sequenced total RNA from 18 OS bone samples (paired normal-tumor biopsies). We found statistically significant (FDR <0.05) 26 differentially expressed transcript variants of LEPROT gene with different expressions in normal and tumor samples. These findings contribute to the understanding of molecular mechanisms of OS development and provide encouragement to pursue further research.

The Genetic Variations Associated With Time to Aseptic Loosening After Total Joint Arthroplasty.

BACKGROUND: Total joint arthroplasty (TJA) is one of the most frequent surgical procedures performed in modern hospitals, and aseptic loosening is the most common indication for revision surgeries. We conducted a systemic exploration of potential genetic determinants for early aseptic loosening. METHODS: Data from 423 patients undergoing TJA were collected and analyzed. Three analytical groups were formed based on joint arthroplasty status. Group 1 were TJA patients without symptoms of aseptic loosening of at least 1 year, group 2 were patients with primary TJA, and group 3 were patients receiving revision surgery because of aseptic loosening. Genome-wide genotyping comparing genotype frequencies between patients with and without aseptic loosening (group 3 vs groups 1 and 2) was conducted. A case-control association analysis and linear modeling were applied to identify the impact of the identified genes on implant survival with time to the revision as an outcome measure. RESULTS: We identified 52 single-nucleotide polymorphisms (SNPs) with a genome-wide suggestive P value less than 10-5 to be associated with the implant loosening. The most remarkable odds ratios (OR) were found with the variations in the IFIT2/IFIT3 (OR, 21.6), CERK (OR, 12.6), and PAPPA (OR, 14.0) genes. Variations in the genotypes of 4 SNPs-rs115871127, rs16823835, rs13275667, and rs2514486-predicted variability in the time to aseptic loosening. The time to aseptic loosening varied from 8 to 16 years depending on the genotype, indicating a substantial effect of genetic variance. CONCLUSION: Development of the aseptic loosening is associated with several genetic variations and we identified at least 4 SNPs with a significant effect on the time for loosening. These data could help to develop a personalized approach for TJA and loosening management.

Analysis of the Expression of Repetitive DNA Elements in Osteosarcoma.

Osteosarcoma (OS) is a rare malignant bone tumor. It affects mostly young persons and has poor outcome with the present treatment. No improvement was observed since the introduction of chemotherapy. The better understanding of osteosarcoma development could indicate better management strategy. Repetitive DNA elements were found to play a role in cancer mechanism especially in epithelial tumors but not yet analyzed in osteosarcoma. We conducted the study to analyse the expression profile of repetitive elements (RE) in osteosarcoma. Methods: Fresh bone paired (tumor and normal bone) samples were obtained from excised parts of tumors of 18 patients with osteosarcoma. We performed sequencing of RNA extracted from 36 samples (18 tumor tissues and 18 normal bone for controls), mapped raw reads to the human genome and identified the REs. EdgeR package was used to analyse the difference in expression of REs between osteosarcoma and normal bone. Results: 82 REs were found differentially expressed (FDR < 0.05) between osteosarcoma and normal bone. Out of all significantly changed REs, 35 were upregulated and 47 were downregulated. HERVs (THE1C-int, LTR5, MER57F and MER87B) and satellite elements (HSATII, ALR-alpha) were the most significantly differential expressed elements between osteosarcoma and normal tissues. These results suggest significant impact of REs in the osteosarcoma. The role of REs should be further studied to understand the mechanism they have in the genesis of osteosarcoma.

Whole transcriptome analysis identifies differentially regulated networks between osteosarcoma and normal bone samples.

We performed whole transcriptome analysis of osteosarcoma bone samples. Initially, we sequenced total RNA from 36 fresh-frozen samples (18 tumoral bone samples and 18 non-tumoral paired samples) matching in pairs for each osteosarcoma patient. We also performed independent gene expression analysis of formalin-fixed paraffin-embedded samples to verify the RNAseq results. Formalin-fixed paraffin-embedded samples allowed us to analyze the effect of chemotherapy. Data were analyzed with DESeq2, edgeR and Reactome packages of R. We found 5365 genes expressed differentially between the normal bone and osteosarcoma tissues with an FDR below 0.05, of which 3399 genes were upregulated and 1966 were downregulated. Among those genes, BTNL9, MMP14, ABCA10, ACACB, COL11A1, and PKM2 were expressed differentially with the highest significance between tumor and normal bone. Functional annotation with the reactome identified significant changes in the pathways related to the extracellular matrix degradation and collagen biosynthesis. It was suggested that chemotherapy may induce the modification of ECM with important collagen biosynthesis. Taken together, our results indicate that changes in the degradation of extracellular matrix seem to be an important mechanism of osteosarcoma and efficient chemotherapy induces the genes related to bone formation. Impact statement Osteosarcoma is a rare disease but it is of interest to many scientists all over the world because the current standard treatment still has poor results. We sequenced total RNA from 36 fresh-frozen paired samples (18 tumoral bone samples and 18 non-tumoral paired samples) from osteosarcoma patients. We found that differences in the gene expressions between the normal and affected bones reflected the changes in the regulation of the degradation of collagen and extracellular matrix. We believe that these findings contribute to the understanding of OS and suggest ideas for further studies.

Body composition and inflammatory markers in pubertal girls: Comparison between athletes and non-athletic controls.

Various inflammation parameters are increased with childhood obesity, but few comparable data are found in lean growing athletes. This study aims to characterize differences in 12 simultaneously measured inflammatory parameters between pubertal rhythmic gymnasts (RG) and untrained controls (UC), and to examine the relationship between body composition and inflammatory markers. Sixty 10-12-year-old girls were divided into RG (n = 30) and UC (n = 30). Fat mass (FM) and fat free mass (FFM) were measured by dual-energy X-ray absorptiometry. Leptin and 12 inflammatory parameters (interleukin [IL]-2, IL-4, IL-6, IL-8, IL-10, vascular endothelial growth factor, interferon-gamma [IFN-γ], tumor necrosis factor-alpha, IL-1α, IL-1ß, monocyte chemotactic protein-1 and epidermal growth factor) were measured from fasting blood samples. No differences were seen in 12 inflammatory markers between studied groups. As expected, leptin (RG: 2.4 ± 1.1; UC: 7.6 ± 4.2 ng ml-1) and FM (RG: 7.3 ± 2.3; UC: 11.8 ± 5.1 kg) were lower (p < .05) in RG compared to UC. In the whole group of lean pubertal girls, 69.0% of the variability in body FM was determined by leptin, and 11.2% of the variability in body FFM was explained by IFN-γ. In conclusion, measured 12 inflammatory biomarkers were not different between RG and UC, despite lower leptin and FM in RG. In lean pubertal girls, IFN-γ was independently associated with FFM, and leptin with FM.

Blocking Tumor-Educated MSC Paracrine Activity Halts Osteosarcoma Progression.

Purpose: Human osteosarcoma is a genetically heterogeneous bone malignancy with poor prognosis despite the employment of aggressive chemotherapy regimens. Because druggable driver mutations have not been established, dissecting the interactions between osteosarcoma cells and supporting stroma may provide insights into novel therapeutic targets.Experimental Design: By using a bioluminescent orthotopic xenograft mouse model of osteosarcoma, we evaluated the effect of tumor extracellular vesicle (EV)-educated mesenchymal stem cells (TEMSC) on osteosarcoma progression. Characterization and functional studies were designed to assess the mechanisms underlying MSC education. Independent series of tissue specimens were analyzed to corroborate the preclinical findings, and the composition of patient serum EVs was analyzed after isolation with size-exclusion chromatography.Results: We show that EVs secreted by highly malignant osteosarcoma cells selectively incorporate a membrane-associated form of TGFß, which induces proinflammatory IL6 production by MSCs. TEMSCs promote tumor growth, accompanied with intratumor STAT3 activation and lung metastasis formation, which was not observed with control MSCs. Importantly, intravenous administration of the anti-IL6 receptor antibody tocilizumab abrogated the tumor-promoting effects of TEMSCs. RNA-seq analysis of human osteosarcoma tissues revealed a distinct TGFß-induced prometastatic gene signature. Tissue microarray immunostaining indicated active STAT3 signaling in human osteosarcoma, consistent with the observations in TEMSC-treated mice. Finally, we isolated pure populations of EVs from serum and demonstrated that circulating levels of EV-associated TGFß are increased in osteosarcoma patients.Conclusions: Collectively, our findings suggest that TEMSCs promote osteosarcoma progression and provide the basis for testing IL6- and TGFß-blocking agents as new therapeutic options for osteosarcoma patients. Clin Cancer Res; 23(14); 3721-33. ©2017 AACR.

The clinical features of osteogenesis imperfecta in Vietnam.

PURPOSE: Osteogenesis imperfecta (OI) has not been studied in a Vietnamese population before. The aim of this study was to systematically collect epidemiological information, investigate clinical features and create a clinical database of OI patients in Vietnam for future research and treatment strategy development. METHOD: Participants underwent clinical and physical examinations; also medical records were reviewed. Genealogical information was collected and family members' phenotypical manifestations recorded. Cases were classified according to the Sillence classification. RESULTS: In total, 146 OI patients from 120 families were studied: 46 with OI Type I, 46 with Type III and 54 with Type IV. Almost patients had skeletal deformations. One hundred and forty-two had a history of fractures, 117 blue sclera, 89 dentinogenesis imperfecta and 26 hearing loss. The total number of fractures was 1,932. Thirty-four patients had intra-uterine fractures and nine had perinatal fractures. Surgery was performed 163 times in 58 patients; 100 osteosyntheses and 63 osteotomies. Bisphosphonate treatment was used in 37 patients. The number of affected individuals and predominance of severe forms of OI indicate that the disease is under diagnosed in Vietnam, especially in cases without a family history or with mild form of OI. Deformities appeared in all patients with different severity and localisation, affecting mostly the lower limbs. OI medical and surgical treatment rates are low and in most cases surgery was performed due to fractures. CONCLUSIONS: Compared to previous studies, our results indicate a lower OI prevalence and greater severity of symptoms in the Vietnamese population when compared with other areas. Further investigation, improved diagnosis and treatment are needed to increase the patients' quality of life.

Longitudinal associations between bone and adipose tissue biochemical markers with bone mineralization in boys during puberty.

BACKGROUND: We investigated longitudinal relationships between the biochemical markers of bone and adipose tissue with bone mineral content (BMC), bone mineral density (BMD), moderate-to-vigorous physical activity (MVPA) and sedentary time (SED) in pubertal boys. METHODS: Ninety-six boys (11.9 ± 0.6 years old) were measured at baseline, after 12 and 24 months. Body composition (fat mass [FM], lean body mass [LBM]), and whole body (WB), lumbar spine (LS) and femoral neck (FN) BMD and BMC were assessed. Additionally, serum leptin, adiponectin, osteocalcin (OC) and C-terminal telopeptide of type I collagen (CTX) were measured. RESULTS: OC had a strong longitudinal inverse effect on changes in WB_BMD (p < 0.001) and LS_BMD (p = 0.021), while CTX had an inverse effect only on changes in FN_BMD (p = 0.011). Leptin had an inverse effect on changes in WB_BMC/WB_BMD (p = 0.001), FN_BMD (p = 0.002) and LS_BMD (p = 0.001). MVPA showed a longitudinal inverse effect on changes in leptin (p = 0.030), however no longitudinal effect of SED to biochemical markers of bone and adipose tissue was found. CONCLUSIONS: Bone metabolism markers have negative effect on bone mineral accrual during puberty. Increases in MVPA affect leptin, suggesting a positive link of MVPA through leptin metabolism on increases in bone mineralization during puberty.

Adipocytokines and bone metabolism markers in relation to bone mineral values in early pubertal boys with different physical activity.

BACKGROUND: We aimed to examine the associations of adipocytokines and circulating bone metabolism markers with bone mineral parameters in early pubertal boys with different physical activity level. METHODS: Eighty-six early pubertal boys were divided into active and non-active boys according to the accumulated moderate-to-vigorous physical activity (MVPA) level. Body composition and bone mineral parameters were assessed and testosterone, leptin, adiponectin, osteocalcin (OC), and C-terminal telopeptide of type I collagen (CTX) were measured. RESULTS: Active subjects had significantly lower (p<0.05) body mass, body mass index (BMI), fat mass (FM), leptin, and sedentary time values, while non-active subjects had lower (p<0.05) vigorous physical activity level and femoral neck bone mineral density (FN-BMD). OC contributed to the models in physically active group and explained 6.6% and 9.7% of variance in whole body (WB) [F(5,44)=10.847; p<0.001] and lumbar spine bone mineral content (LS-BMC) [F(5,44)=4.158; p=0.004], respectively. No other biochemical parameters were found to be related to bone mineral parameters in either the active or non-active group. CONCLUSIONS: Bone metabolism markers were positively correlated with bone mineral values only in active pubertal boys. Leptin and adiponectin were not related to bone mineral parameters in active and non-active pubertal boys.

Laminin-rich blood vessels display activated growth factor signaling and act as the proliferation centers in Dupuytren's contracture.

INTRODUCTION: Dupuytren's contracture (DC) is a chronic fibroproliferative disease of the hand, which is characterized by uncontrolled proliferation of atypical myofibroblasts at the cellular level. We hypothesized that specific areas of the DC tissue are sustaining the cell proliferation and studied the potential molecular determinants that might contribute to the formation of such niches. METHODS: We studied the expression pattern of cell proliferation marker Ki67, phosphorylated AKT (Ak mouse strain thymoma) kinase, DC-associated growth factors (connective tissue growth factor (CTGF), basic fibroblast growth factor (bFGF), insulin-like growth factor 2 (IGF-2)) and extracellular matrix components (laminins, fibronectin, collagen IV) in DC tissue and normal palmar fascia using immunofluorescence microscopy and quantitative real-time polymerase chain reaction (qPCR). RESULTS: We found that proliferative cells in the DC nodules were concentrated in the immediate vicinity of small blood vessels and localized predominantly in the myofibroblast layer. Correspondingly, the DC-associated blood vessels contained increased levels of phosphorylated AKT, a hallmark of activated growth factor signaling. When studying the expression of potential activators of AKT signaling we found that the expression of bFGF was confined to the endothelium of the small blood vessels, IGF-2 was present uniformly in the DC tissue and CTGF was expressed in the DC-associated sweat gland acini. In addition, the blood vessels in DC nodules contained increased amounts of laminins 511 and 521, which have been previously shown to promote the proliferation and stem cell properties of different cell types. CONCLUSIONS: Based on our findings, we propose that in the DC-associated small blood vessels the presence of growth factors in combination with favorable extracellular matrix composition provide a supportive environment for sustained proliferation of myofibroblasts and thus the blood vessels play an important role in DC pathogenesis.

Whole exome sequencing of a single osteosarcoma case--integrative analysis with whole transcriptome RNA-seq data.

BACKGROUND: Osteosarcoma (OS) is a prevalent primary malignant bone tumour with unknown etiology. These highly metastasizing tumours are among the most frequent causes of cancer-related deaths. Thus, there is an urgent need for different markers, and with our study, we were aiming towards finding novel biomarkers for OS. METHODS: For that, we analysed the whole exome of the tumorous and non-tumour bone tissue from the same patient with OS applying next-generation sequencing. For data analysis, we used several softwares and combined the exome data with RNA-seq data from our previous study. RESULTS: In the tumour exome, we found wide genomic rearrangements, which should qualify as chromotripsis-we detected almost 3,000 somatic single nucleotide variants (SNVs) and small indels and more than 2,000 copy number variants (CNVs) in different chromosomes. Furthermore, the somatic changes seem to be associated to bone tumours, whereas germline mutations to cancer in general. We confirmed the previous findings that the most significant pathway involved in OS pathogenesis is probably the WNT/ß-catenin signalling pathway. Also, the IGF1/IGF2 and IGF1R homodimer signalling and TP53 (including downstream tumour suppressor gene EI24) pathways may have a role. Additionally, the mucin family genes, especially MUC4 and cell cycle controlling gene CDC27 may be considered as potential biomarkers for OS. CONCLUSIONS: The genes, in which the mutations were detected, may be considered as targets for finding biomarkers for OS. As the study is based on a single case and only DNA and RNA analysis, further confirmative studies are required.

Negative correlation between serum IL-6 level and cardiorespiratory fitness in 10- to 11-year-old boys with increased BMI.

BACKGROUND/AIMS: Some markers of inflammation have been found to be associated with cardiorespiratory fitness levels, but only few studies have studied this in overweight children. The aim of this study was to investigate associations between markers of inflammation and the fitness levels measured by peak oxygen consumption (VO(2peak) and VO(2peak)/kg) in boys with increased body mass index (BMI) and with normal BMI. PARTICIPANTS/METHODS: Subjects were 38 boys with BMI above 85th percentile (OWB) and 38 boys with normal BMI (NWB) at the age of 10 to 11 years. Serum concentrations of IL-2, IL-4, IL-6, IL-8, IL-10, VEGF, IFNγ, TNF-α, IL-1α, IL-1ß, MCP-1, EGF, CRP and associations with measured cardiorespiratory fitness levels were studied. High-sensitive chips were used to measure 13 markers of inflammation. RESULTS: Mean VO(2peak) was significantly higher (2.1±0.3 vs. 1.8±0.3 L/min; p<0.05) and mean VO(2peak)/kg significantly lower (33.7±4.7 vs. 48.9±6.4 mL/min/kg; p<0.05) in OWB than in NWB group. Out of 13 measured biochemical markers IL-6 correlated with VO(2peak)/kg (r=-0.37; p<0.05) and TNF-α with VO(2peak) (r=0.41; p<0.01) in OWB. BMI and IL-6 together explained 44.5% of the variability of VO(2peak)/kg in the OWB group. CONCLUSIONS: Overweight boys had lower cardiorespiratory fitness level measured by VO(2peak)/kg and this was negatively correlated with serum IL-6 level. Measurement of serum IL-6 level in overweight boys may help to identify subjects who need specific exercise formats to achieve maximal beneficial health effects and to reduce their risk for the development of type 2 diabetes and atherosclerosis later in life.

Effects of denosumab on bone histomorphometry: the FREEDOM and STAND studies.

Denosumab, a human monoclonal antibody against RANKL, reversibly inhibits osteoclast-mediated bone resorption and has been developed for use in osteoporosis. Its effects on bone histomorphometry have not been described previously. Iliac crest bone biopsies were collected at 24 and/or 36 months from osteoporotic postmenopausal women in the FREEDOM study (45 women receiving placebo and 47 denosumab) and at 12 months from postmenopausal women previously treated with alendronate in the STAND study (21 continuing alendronate and 15 changed to denosumab at trial entry). Qualitative histologic evaluation of biopsies was unremarkable. In the FREEDOM study, median eroded surface was reduced by more than 80% and osteoclasts were absent from more than 50% of biopsies in the denosumab group. Double labeling in trabecular bone was observed in 94% of placebo bones and in 19% of those treated with denosumab. Median bone-formation rate was reduced by 97%. Among denosumab-treated subjects, those with double labels and those with absent labels had similar levels of biochemical markers of bone turnover. In the STAND trial, indices of bone turnover tended to be lower in the denosumab group than in the alendronate group. Double labeling in trabecular bone was seen in 20% of the denosumab biopsies and in 90% of the alendronate samples. Denosumab markedly reduces bone turnover and also reduces fracture numbers. Longer follow-up is necessary to determine how long such low turnover is safe.