Efficacy and safety of postoperative single-dose anticoagulant treatment in preventing venous thromboembolism after revision hip arthroplasty: a retrospective cohort study.

PURPOSE: The purpose of the study is to determine the efficacy and safety of postoperative single-dose anticoagulant treatment in preventing venous thromboembolism (VTE) after revision THA, in comparison with a multiple-dose chemoprophylaxis protocol. METHODS: We retrospectively compared 295 patients undergoing revision THA who received multiple-dose chemoprophylaxis protocol (40 mg low-molecular-weight heparin once and oral rivaroxaban for 10 days) or single-dose chemoprophylaxis protocol (40 mg low-molecular-weight heparin once) for VTE. The patients in both groups performed active lower limb exercises. Each group was further stratified into subgroups based on the aetiology of revision. The incidence of VTE, wound complications within three months, hidden blood loss (HBL), transfusion rate, and surgical drainage duration were recorded. RESULTS: The incidence rates of VTE (P = 0.870) did not differ between the two prophylaxis protocols. However, significant differences were observed in wound complications within three months (P = 0.002), HBL (P = 0.015), transfusion rate (P = 0.028). Surgical drainage duration was also shorter in the single-dose chemoprophylaxis group (P = 0.0023). In the subgroup analysis, the use of single-dose chemoprophylaxis protocol cannot significantly reduce HBL and transfusion rate after septic revision THA. The use of multiple-dose chemoprophylaxis protocol (OR = 2.89, P = 0.002) and high BMI (OR = 1.09, P = 0.037) were independent risk factors of wound complications. CONCLUSIONS: Single-dose chemoprophylaxis protocol effectively and safely prevented VTE after revision THA compared with multiple-dose chemoprophylaxis protocol. The effect in reducing HBL and postoperative transfusion rate was limited in septic revision.

Single-stage revision for chronic periprosthetic joint infection after knee and hip arthroplasties: indications and treatments.

Single-stage revision for chronic periprosthetic joint infection has been introduced 40 years ago. This option is gaining more and more attention as well as popularity. It is a reliable treatment for the chronic periprosthetic joint infection after knee and hip arthroplasties when implemented by an experienced multi-disciplinary team. However, its indications and corresponding treatments remain controversial. This review focused on the indications and specific treatments related to the option, with an attempt to help surgeons to use this method with more favorable outcomes.

Midterm Survival of a Varus-Valgus Constrained Implant following One-Stage Revision for Periprosthetic Joint Infection: A Single-Center Study.

Revision total knee arthroplasty (TKA) is challenging to perform in patients with periprosthetic joint infection (PJI) due to the difficulty of eradicating the infection and potential for bone and ligamentous deficits. The current study aimed to evaluate the midterm survival of varus-valgus constrained (VVC) implants used in one-stage revision TKA for chronic PJI at our institution. This retrospective analysis included 132 patients with chronic PJI who underwent one-stage revision using a VVC implant. The average follow-up was 51.6 months (range: 24-85 months). Five-year survival analysis was performed to set recurrent infection and mechanical failure as the end point. Hospital for special surgery (HSS) as functional outcomes was evaluated preoperatively and at the latest follow-up. A total of 12 patients (9.1%) underwent retreatment for reinfection (nine patients) and mechanical failure (three patients). The overall 5-year revision-free survival was 82.7%, the infection-free survival was 91.1%, and the mechanical failure-free survival was 98.3%. The preoperative HSS knee score improved from 35.6 points (range: 24.3-47.7 points) preoperatively to 76.8 points (range: 57.2-87.6 points) at the latest follow-up. Complications were identified in 20 patients (15.2%) which included aseptic osteolysis in 4 cases, acceptable flexion instability in 3 cases, arthrofibrosis in 2 patients, hematomas in 2 cases, calf intermuscular venous thrombosis in 6 patients, and femoral stem tip pain in 3 cases. This is the first study to report the outcomes of one-stage revision using VVC implants for knee PJI. Improved functional outcomes and good midterm survival are demonstrated at an average follow-up of 51.6 months.

Deep insights into the viscosity of deep eutectic solvents by an XGBoost-based model plus SHapley Additive exPlanation.

Deep eutectic solvents (DESs) are emerging as novel green solvents for the processes of mass transport and heat transfer, in which the viscosity of DESs is important for their industrial applications. However, for DESs, the measurement of viscosity is time-consuming, and there are many factors influencing the viscosity, which impedes their wider application. This study aims to develop a data-driven model which could accurately and rapidly predict the viscosity of diverse DESs at different temperatures, and furthermore boost the design and screening of novel DESs. In this work, we collected 107 DESs with 994 experimental values of viscosity from published works. Given the significant effect of water on viscosity, the water content of each collected DES was labeled. The Morgan fingerprint was first employed as a feature to describe the chemical environment of DESs. And four machine learning algorithms were used to train models: support vector regression (SVR), random forest (RF), neural network (NN), and extreme gradient boosting (XGBoost), and XGBoost showed the best predictive performance. In combination with the powerful interpretation method SHapley Additive exPlanation (SHAP), we further revealed the positive or negative effect of features on viscosity. Overall, this work provides a machine learning model which could predict viscosity precisely and facilitate the design and application of DESs.

Effects of intra-articular D-amino acids combined with systemic vancomycin on an experimental Staphylococcus aureus-induced periprosthetic joint infection.

BACKGROUND: The D-isoforms of amino acids (D-AAs) exhibit anti-biofilm potential against a diverse range of bacterial species in vitro, while its role in vivo remains unclear. The aim of this study was to investigate the effects of a combination of D-AAs and vancomycin on a PJI rat model. METHODS: Eight-week-old male SD rats were randomized to the control group, sham group, vancomycin group, D-AAs-vancomycin group. After treatment for 6 weeks, we analysed the levels of inflammatory factors in serum, behavioural change, imaging manifestations. The anti-biofilm ability of D-AAs was detected by crystal violet staining and scanning electron microscope observation, and its ability to assist antibiotics in killing bacteria was assessed by culture of bacteria. Additionally, micro-CT and histological analysis were used to evaluate the impact of D-AAs combined with vancomycin on the bone remodelling around the prosthesis. RESULTS: The group treated with a D-AAs-vancomycin combination sustained normal weight gain and exhibited reduced the serum levels of α2M, IL-1ß, IL-6, IL-10, TNF-α and PGE2. Moreover, treated with D-AAs in combination with vancomycin improved the weight-bearing activity performance, increased the sizes and widths of distal femurs, and improved Rissing scale scoring. In particular, treatment using D-AAs enhanced the ability of vancomycin to eradicate Staphylococcus aureus, as demonstrated by the dispersion of existing biofilms and the inhibition of biofilm formation that occurred in a concentration-dependent manner. This treatment combination also resulted in a reduction in bacterial burden with in the soft tissues, bones, and implants. Furthermore, D-AAs-vancomycin combination treatment attenuated abnormal bone remodelling around the implant, as evidenced by an observed increase in BMD, BV/TV, and Tb.Th and the presence of reduced Trap+ osteoclasts and elevated osterix+ osteo-progenitors. CONCLUSIONS: Combining D-AAs with vancomycin provides an effective therapeutic strategy for the treatment of PJI by promoting biofilm dispersion to enhance antimicrobial activity.

Evaluation of phenotype-driven gene prioritization methods for Mendelian diseases.

It's challenging work to identify disease-causing genes from the next-generation sequencing (NGS) data of patients with Mendelian disorders. To improve this situation, researchers have developed many phenotype-driven gene prioritization methods using a patient's genotype and phenotype information, or phenotype information only as input to rank the candidate's pathogenic genes. Evaluations of these ranking methods provide practitioners with convenience for choosing an appropriate tool for their workflows, but retrospective benchmarks are underpowered to provide statistically significant results in their attempt to differentiate. In this research, the performance of ten recognized causal-gene prioritization methods was benchmarked using 305 cases from the Deciphering Developmental Disorders (DDD) project and 209 in-house cases via a relatively unbiased methodology. The evaluation results show that methods using Human Phenotype Ontology (HPO) terms and Variant Call Format (VCF) files as input achieved better overall performance than those using phenotypic data alone. Besides, LIRICAL and AMELIE, two of the best methods in our benchmark experiments, complement each other in cases with the causal genes ranked highly, suggesting a possible integrative approach to further enhance the diagnostic efficiency. Our benchmarking provides valuable reference information to the computer-assisted rapid diagnosis in Mendelian diseases and sheds some light on the potential direction of future improvement on disease-causing gene prioritization methods.

Mid-Term Outcomes of Cementless Total Hip Arthroplasty in Adult Patients with Childhood Hip Infection.

BACKGROUND Patients with hip joint infections in childhood often have many aftereffects of different degrees, regardless of the kind of treatment or natural course. Total hip arthroplasty is currently the most effective treatment for sequelae of childhood hip septic or tuberculous infection. This is a mid-term follow-up study of treatment results of patients who had undergone total hip arthroplasty (THA) with cementless prostheses. MATERIAL AND METHODS We retrospectively analyzed and followed 45 patients (45 hips) who underwent THA with cementless prostheses between 2010 and 2017. There were 45 patients, including 17 men and 28 women. The average age of the patients was 46 years (range, 18-67 years). All hip infections occurred in early childhood or adolescence, and the mean interval between initial infection and THA was 38.2 years (range, 15-60 years). The mean follow-up was 6.1 years (range, 2.7-9.5 years). RESULTS Two patients underwent revision surgery because of loosening of the prosthesis, and 1 patient underwent revision surgery because of a new infection with no relationship with childhood infection during the follow-up. The average Harris hip scores significantly increased from 43.1 to 86.4 (P<0.01), and the average visual analog scale significantly increased from 4.6 to 1.7 (P<0.01). The hip dysfunction and osteoarthritis outcome scores were also significantly changed (P<0.01) at the final follow-up. There were 2 cases of transient sciatic nerve palsy and intraoperative periprosthetic fractures in 3 cases. During follow-up, single revision was performed after 6 years of primary arthroplasty because of aseptic loosening in 2 cases and prosthesis infection in 1 case, which was not related to childhood pathogens. CONCLUSIONS THA for patients with sequelae of hip joint infection has a satisfactory effect that can effectively relieve joint pain and improve hip function. The recurrence rate of infection after either pyogenic infection or tuberculous is very low. The mid-term outcomes of THA in this setting were satisfactory, with high prosthesis survivorship and hip function scores.

Premigratory neural crest stem cells generate enteric neurons populating the mouse colon and regulating peristalsis in tissue-engineered intestine.

Hirschsprung's disease (HSCR) is a common congenital defect. It occurs when bowel colonization by neural crest-derived enteric nervous system (ENS) precursors is incomplete during the first trimester of pregnancy. Several sources of candidate cells have been previously studied for their capacity to regenerate the ENS, including enteric neural crest stem cells (En-NCSCs) derived from native intestine or those simulated from human pluripotent stem cells (hPSCs). However, it is not yet known whether the native NCSCs other than En-NCSCs would have the potential of regenerating functional enteric neurons and producing neuron dependent motility under the intestinal environment. The present study was designed to determine whether premigratory NCSCs (pNCSCs), as a type of the nonenteric NCSCs, could form enteric neurons and mediate the motility. pNCSCs were firstly transplanted into the colon of adult mice, and were found to survive, migrate, differentiate into enteric neurons, and successfully integrate into the adult mouse colon. When the mixture of pNCSCs and human intestinal organoids was implanted into the subrenal capsule of nude mice and grown into the mature tissue-engineered intestine (TEI), the pNCSCs-derived neurons mediated neuron-dependent peristalsis of TEI. These results show that the pNCSCs that were previously assumed to not be induced by intestinal environment or cues can innervate the intestine and establish neuron-dependent motility. Future cell candidates for ENS regeneration may include nonenteric NCSCs.

Multicup reconstruction technique for the management of severe protrusio acetabular defects.

BACKGROUND: In revision hip arthroplasty, managing the large protrusio acetabular defects remains a challenge. The report described a novel technique which employs a trabecular metal revision shell as a super-augment to buttress the superior medial structure. METHODS: Between January 2015 and December 2018, the multicup reconstruction was performed in 21 patients with severe protrusio acetabular defects. The revision shell, plus two similar porous acetabular components was implanted into the initial shell to create a "multicup" construct. The functional outcomes were evaluated in terms of the Harris Hip Score. Acetabular loosening, restoration of hip center of rotation, and bone ingrowth etc., were radiographically assessed. The survival rate of the implants was also evaluated. RESULTS: A followup lasting a mean time of 31 months (range, 18-57 months) revealed that the average Harris Hip Score improved from preoperative 37.0 ± 7.1 to postoperative 76.4 ± 9.0. There were no revisions due to acetabular loosening. The horizontal offset increased by an average of 14 mm, and the vertical offset decreased by an average of 18 mm. Eighteen of the 21 patients (86 %) met at least 3 of 5 criteria associated with bone ingrowth. The survivorship free from re-revision for acetabular loosening after 2 years was 100 %. CONCLUSIONS: The multicup reconstruction technique was a simplified re-revision procedure for managing the severe protrusio acetabular defects and could achieve a high survival rate. LEVEL OF EVIDENCE: Therapeutic study, Level IVa.

Outcome of Irrigation and Debridement With Topical Antibiotics Delivery for the Management of Periprosthetic Joint Infection Occurring Within 3 Months Since the Primary Total Joint Arthroplasty.

BACKGROUND: Irrigation and debridement with modular component exchange is appealing for surgeons to treat early-stage periprosthetic joint infection (PJI). However, the indication, perioperative protocol, and success rate remain controversial. This study is the first one to present results of debridement, antibiotics, and implant retention (DAIR) with integrated MIT (modular component exchange, povidone-iodine and topical antibiotics delivery) protocol for treating PJI occurring within 3 months since the primary total joint arthroplasty. METHODS: We retrospectively analyzed patients who received DAIR with MIT protocol in our department between January 2011 and May 2018. Topical antibiotics were delivered in all cases. Topical antibiotics infusion was applied for those infected with multidrug-resistant bacteria, fungus, polymicrobial infection, and culture negative one. Failure was defined as additional surgical intervention for infection after DAIR; persistent sinus tract, drainage or excessive joint pain; need for suppressive antibiotics therapy due to the infection; infection relapse with the same pathogen; reinfection with different microorganism; and infection-related death. RESULTS: A total of 73 patients with a mean age of 63.30 ± 10.97 years were included in this study, including 43 men and 30 women. There are 41 knees and 32 hips. Thirty patients had sinus tract. With a mean follow-up of 63.79 ± 18.57 months, there were 9 failures in total with an overall success rate of 87.67%. The success rate was 88.57% and 86.84% for those receiving topical antibiotics infusion postoperatively and those without. CONCLUSIONS: DAIR with a standard MIT protocol is a viable and safe option for PJI occurring within 3 months since the primary total joint arthroplasty. LEVEL OF EVIDENCE: Level 4, therapeutic study.

Effective treatment of single-stage revision using intra-articular antibiotic infusion for culture-negative prosthetic joint infection.

AIMS: In the absence of an identified organism, single-stage revision is contraindicated in prosthetic joint infection (PJI). However, no studies have examined the use of intra-articular antibiotics in combination with single-stage revision in these cases. In this study, we present the results of single-stage revision using intra-articular antibiotic infusion for treating culture-negative (CN) PJI. METHODS: A retrospective analysis between 2009 and 2016 included 51 patients with CN PJI who underwent single-stage revision using intra-articular antibiotic infusion; these were compared with 192 culture-positive (CP) patients. CN patients were treated according to a protocol including intravenous vancomycin and a direct intra-articular infusion of imipenem and vancomycin alternately used in the morning and afternoon. In the CP patients, pathogen-sensitive intravenous (IV) antibiotics were administered for a mean of 16 days (12 to 21), and for resistant cases, additional intra-articular antibiotics were used. The infection healing rate, Harris Hip Score (HHS), and Hospital for Special Surgery (HSS) knee score were compared between CN and CP groups. RESULTS: Of 51 CN patients, 46 (90.2%) required no additional medical treatment for recurrent infection at a mean of 53.2 months (24 to 72) of follow-up. Impaired kidney function occurred in two patients, and one patient had a local skin rash. No significant difference in the infection control rate was observed between CN and CP PJIs (90.2% (46/51) versus 94.3% (181/192); p = 0.297). The HHS of the CN group showed no substantial difference from that of CP cases (79 versus 81; p = 0.359). However, the CN group showed a mean HSS inferior to that of the CP group (76 versus 80; p = 0.027). CONCLUSION: Single-stage revision with direct intra-articular antibiotic infusion can be effective in treating CN PJI, and can achieve an infection control rate similar to that in CP patients. However, in view of systemic toxicity, local adverse reactions, and higher costs, additional strong evidence is needed to verify these treatment regimens. Cite this article: Bone Joint J 2020;102-B(3):336-344.

Artesunate alleviates interleukin­1ß­induced inflammatory response and apoptosis by inhibiting the NF­κB signaling pathway in chondrocyte­like ATDC5 cells, and delays the progression of osteoarthritis in a mouse model.

Osteoarthritis (OA) is a progressive and degenerative joint disorder that is highly prevalent worldwide and for which there is currently no effective medical therapy. Artesunate (ART), a natural compound used to treat malaria, possesses diverse biological properties, including the regulation of inflammation and apoptosis in various cells; however, its role in OA remains unclear. The aim of the present study was to investigate the effects of ART on interleukin (IL)­1ß­induced chondrocyte­like ATDC5 cells and in an OA mouse model. The results revealed that ART dose­dependently relieved the inhibitory effect of IL­1ß on cell viability. Moreover, ART significantly reduced the overexpression of matrix metalloproteinase (MMP)­3, MMP­13, a disintegrin and metalloproteinase with thrombospondin motifs­5 and cyclooxygenase­2 at both the gene and protein levels in chondrocyte­like ATDC5 cells stimulated by IL­1ß. Furthermore, ART decreased the expression of pro­apoptotic Bax, cleaved caspase­3 and cleaved caspase­7 in a dose­dependent manner, and increased the expression of the anti­apoptotic factor Bcl­2. These changes were mediated by the inhibitory effect of ART on the nuclear factor­κB signaling pathway, defined as repression of the phosphorylation of IκBα and p65, and improved redistribution of p65. Additionally, ART blocked the advancement of the calcified cartilage zone and the loss of proteoglycan, and lowered histological scoring of OA in a mouse model. Taken together, these results indicate that ART may be of value as a therapeutic agent for OA.

Artesunate, an Anti-Malaria Agent, Attenuates Experimental Osteoarthritis by Inhibiting Bone Resorption and CD31hiEmcnhi Vessel Formation in Subchondral Bone.

Osteoarthritis (OA) is a common and debilitating joint disease worldwide without interventions available to reverse its progression. Artesunate (ART), an anti-malaria agent, possesses diverse biological activities, including the inhibition of osteoclastogenesis and angiogenesis in various cells, but its role in subchondral bone during OA progression is not known. Here, we explored the curative effects of ART on the pathogenesis of OA in anterior cruciate ligament transection (ACLT) mice models. We found that ART attenuated articular cartilage degeneration, defined by lowered histologic scoring of OA and retarded calcification of the cartilage zone. Moreover, ART improved the expression of lubricin and aggrecan and reduced the expression of collagen X (Col X) and matrix metalloproteinase-13 (MMP-13). In parallel, ART normalized abnormal subchondral bone remodeling by maintaining bone volume fraction (BV/TV) and subchondral bone plate thickness (SBP Th) and reducing trabecular pattern factor (Tb.pf) compared to the vehicle-treated mice. Our results indicated that ART suppressed osteoclastic bone resorption through regulating RANKL-OPG system, restored coupled bone remodeling by indirectly inhibiting TGF-ß/Smad2/3 signaling. Additionally, ART abrogated CD31hiEmcnhi vessel formation via downregulating the expression of vascular endothelial growth factor (VEGF) and angiogenin-1 in subchondral bone. In conclusion, ART attenuates ACLT-induced OA by blocking bone resorption and CD31hiEmcnhi vessel formation in subchondral bone, indicating that this may be a new therapeutic alternative for OA.

Correction: DNA breakpoint assay reveals a majority of gross duplications occur in tandem reducing VUS classifications in breast cancer predisposition genes.

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Association of Breast and Ovarian Cancers With Predisposition Genes Identified by Large-Scale Sequencing.

Importance: Since the discovery of BRCA1 and BRCA2, multiple high- and moderate-penetrance genes have been reported as risk factors for hereditary breast cancer, ovarian cancer, or both; however, it is unclear whether these findings represent the complete genetic landscape of these cancers. Systematic investigation of the genetic contributions to breast and ovarian cancers is needed to confirm these findings and explore potentially new associations. Objective: To confirm reported and identify additional predisposition genes for breast or ovarian cancer. Design, Setting, and Participants: In this sample of 11 416 patients with clinical features of breast cancer, ovarian cancer, or both who were referred for genetic testing from 1200 hospitals and clinics across the United States and of 3988 controls who were referred for genetic testing for noncancer conditions between 2014 and 2015, whole-exome sequencing was conducted and gene-phenotype associations were examined. Case-control analyses using the Genome Aggregation Database as a set of reference controls were also conducted. Main Outcomes and Measures: Breast cancer risk associated with pathogenic variants among 625 cancer predisposition genes; association of identified predisposition breast or ovarian cancer genes with the breast cancer subtypes invasive ductal, invasive lobular, hormone receptor-positive, hormone receptor-negative, and male, and with early-onset disease. Results: Of 9639 patients with breast cancer, 3960 (41.1%) were early-onset cases (≤45 years at diagnosis) and 123 (1.3%) were male, with men having an older age at diagnosis than women (mean [SD] age, 61.8 [12.8] vs 48.6 [11.4] years). Of 2051 women with ovarian cancer, 445 (21.7%) received a diagnosis at 45 years or younger. Enrichment of pathogenic variants were identified in 4 non-BRCA genes associated with breast cancer risk: ATM (odds ratio [OR], 2.97; 95% CI, 1.67-5.68), CHEK2 (OR, 2.19; 95% CI, 1.40-3.56), PALB2 (OR, 5.53; 95% CI, 2.24-17.65), and MSH6 (OR, 2.59; 95% CI, 1.35-5.44). Increased risk for ovarian cancer was associated with 4 genes: MSH6 (OR, 4.16; 95% CI, 1.95-9.47), RAD51C (OR, not estimable; false-discovery rate-corrected P = .004), TP53 (OR, 18.50; 95% CI, 2.56-808.10), and ATM (OR, 2.85; 95% CI, 1.30-6.32). Neither the MRN complex genes nor CDKN2A was associated with increased breast or ovarian cancer risk. The findings also do not support previously reported breast cancer associations with the ovarian cancer susceptibility genes BRIP1, RAD51C, and RAD51D, or mismatch repair genes MSH2 and PMS2. Conclusions and Relevance: The results of this large-scale exome sequencing of patients and controls shed light on both well-established and controversial non-BRCA predisposition gene associations with breast or ovarian cancer reported to date and may implicate additional breast or ovarian cancer susceptibility gene candidates involved in DNA repair and genomic maintenance.

DRG-Derived Neural Progenitors Differentiate into Functional Enteric Neurons Following Transplantation in the Postnatal Colon.

Cell therapy has great promise for treating gastrointestinal motility disorders caused by intestinal nervous system (ENS) diseases. However, appropriate sources, other than enteric neural stem cells and human embryonic stem cells, are seldom reported. Here, we show that neural progenitors derived from the dorsal root ganglion (DRG) of EGFP mice survived, differentiated into enteric neurons and glia cells, migrated widely from the site of injection, and established neuron-muscle connections following transplantation into the distal colon of postnatal mice. The exogenous EGFP+ neurons were physiologically functional as shown by the activity of calcium imaging. This study shows that that other tissues besides the postnatal bowel harbor neural crest stem cells or neural progenitors that have the potential to differentiate into functional enteric neurons in vivo and can potentially be used for intestinal nerve regeneration. These DRG-derived neural progenitor cells may be a choice for cell therapy of ENS disease as an allograft. The new knowledge provided by our study is important for the development of neural crest stem cell and cell therapy for the treatment of intestinal neuropathy.

DNA breakpoint assay reveals a majority of gross duplications occur in tandem reducing VUS classifications in breast cancer predisposition genes.

PURPOSE: Gross duplications are ambiguous in terms of clinical interpretation due to the limitations of the detection methods that cannot infer their context, namely, whether they occur in tandem or are duplicated and inserted elsewhere in the genome. We investigated the proportion of gross duplications occurring in tandem in breast cancer predisposition genes with the intent of informing their classifications. METHODS: The DNA breakpoint assay (DBA) is a custom, paired-end, next-generation sequencing (NGS) method designed to capture and detect deep-intronic DNA breakpoints in gross duplications in BRCA1, BRCA2, ATM, CDH1, PALB2, and CHEK2. RESULTS: DBA allowed us to ascertain breakpoints for 44 unique gross duplications from 147 probands. We determined that the duplications occurred in tandem in 114 (78%) carriers from this cohort, while the remainder have unknown tandem status. Among the tandem gross duplications that were eligible for reclassification, 95% of them were upgraded to pathogenic. CONCLUSION: DBA is a novel, high-throughput, NGS-based method that informs the tandem status, and thereby the classification of, gross duplications. This method revealed that most gross duplications in the investigated genes occurred in tandem and resulted in a pathogenic classification, which helps to secure the necessary treatment options for their carriers.

Detection of structural variation using target captured next-generation sequencing data for genetic diagnostic testing.

PURPOSE: Structural variation (SV) is associated with inherited diseases. Next-generation sequencing (NGS) is an efficient method for SV detection because of its high-throughput, low cost, and base-pair resolution. However, due to lack of standard NGS protocols and a limited number of clinical samples with pathogenic SVs, comprehensive standards for SV detection, interpretation, and reporting are to be established. METHODS: We performed SV assessment on 60,000 clinical samples tested with hereditary cancer NGS panels spanning 48 genes. To evaluate NGS results, NGS and orthogonal methods were used separately in a blinded fashion for SV detection in all samples. RESULTS: A total of 1,037 SVs in coding sequence (CDS) or untranslated regions (UTRs) and 30,847 SVs in introns were detected and validated. Across all variant types, NGS shows 100% sensitivity and 99.9% specificity. Overall, 64% of CDS/UTR SVs were classified as pathogenic/likely pathogenic, and five deletions/duplications were reclassified as pathogenic using breakpoint information from NGS. CONCLUSION: The SVs presented here can be used as a valuable resource for clinical research and diagnostics. The data illustrate NGS as a powerful tool for SV detection. Application of NGS and confirmation technologies in genetic testing ensures delivering accurate and reliable results for diagnosis and patient care.

TumorNext-Lynch-MMR: a comprehensive next generation sequencing assay for the detection of germline and somatic mutations in genes associated with mismatch repair deficiency and Lynch syndrome.

The current algorithm for Lynch syndrome diagnosis is highly complex with multiple steps which can result in an extended time to diagnosis while depleting precious tumor specimens. Here we describe the analytical validation of a custom probe-based NGS tumor panel, TumorNext-Lynch-MMR, which generates a comprehensive genetic profile of both germline and somatic mutations that can accelerate and streamline the time to diagnosis and preserve specimen. TumorNext-Lynch-MMR can detect single nucleotide variants, small insertions and deletions in 39 genes that are frequently mutated in Lynch syndrome and colorectal cancer. Moreover, the panel provides microsatellite instability status and detects loss of heterozygosity in the five Lynch genes; MSH2, MSH6, MLH1, PMS2 and EPCAM. Clinical cases are described that highlight the assays ability to differentiate between somatic and germline mutations, precisely classify variants and resolve discordant cases.

Halofuginone Attenuates Osteoarthritis by Rescuing Bone Remodeling in Subchondral Bone Through Oral Gavage.

Osteoarthritis (OA) is a common debilitating joint disorder worldwide without effective medical therapy. Articular cartilage and subchondral bone act in concert as a functional unit with the onset of OA. Halofuginone is an analog of the alkaloid febrifugine extracted from the plant Dichroa febrifuga, which has been demonstrated to exert inhibition of SMAD 2/3 phosphorylation downstream of the TGF-ß signaling pathway and osteoclastogenesis. To investigate whether halofuginone (HF) alleviates OA after administration by oral gavage, 3-month-old male mice were allocated to the Sham group, vehicle-treated anterior cruciate ligament transection (ACLT) group, and HF-treated ACLT group. The immunostaining analysis indicated that HF reduced the number of matrix metalloproteinase 13 (MMP-13) and collagen X (Col X) positive cells in the articular cartilage. Moreover, HF lowered histologic OA score and prevented articular cartilage degeneration. The micro-computed tomography (µCT) scan showed that HF maintained the subchondral bone microarchitecture, demonstrated by the restoration of bone volume fraction (BV/TV), subchondral bone plate thickness (SBP.Th.), and trabecular pattern factor (Tb.Pf) to a level comparable to that of the Sham group. Immunostaining for CD31 and µCT based angiography showed that the number and volume of vessels in subchondral bone was restored by HF. HF administered by oral gavage recoupled bone remodeling and inhibited aberrant angiogenesis in the subchondral bone, further slowed the progression of OA. Therefore, HF administered by oral gavage could be a potential therapy for OA.