Recruitment of Peroxin 14 to lipid droplets affects lipid storage in Drosophila.

Both peroxisomes and lipid droplets regulate cellular lipid homeostasis. Direct inter-organellar contacts as well as novel roles for proteins associated with peroxisome or lipid droplets occur when cells are induced to liberate fatty acids from lipid droplets. We have shown a non-canonical role for a subset of peroxisome-assembly [Peroxin (Pex)] proteins in this process in Drosophila. Transmembrane proteins Pex3, Pex13 and Pex14 were observed to surround newly formed lipid droplets. Trafficking of Pex14 to lipid droplets was enhanced by loss of Pex19, which directs insertion of transmembrane proteins like Pex14 into the peroxisome bilayer membrane. Accumulation of Pex14 around lipid droplets did not induce changes to peroxisome size or number, and co-recruitment of the remaining Peroxins was not needed to assemble peroxisomes observed. Increasing the relative level of Pex14 surrounding lipid droplets affected the recruitment of Hsl lipase. Fat body-specific reduction of these lipid droplet-associated Peroxins caused a unique effect on larval fat body development and affected their survival on lipid-enriched or minimal diets. This revealed a heretofore unknown function for a subset of Pex proteins in regulating lipid storage. This article has an associated First Person interview with Kazuki Ueda, joint first author of the paper.

A new hypothesis of dinosaur relationships and early dinosaur evolution.

For 130 years, dinosaurs have been divided into two distinct clades-Ornithischia and Saurischia. Here we present a hypothesis for the phylogenetic relationships of the major dinosaurian groups that challenges the current consensus concerning early dinosaur evolution and highlights problematic aspects of current cladistic definitions. Our study has found a sister-group relationship between Ornithischia and Theropoda (united in the new clade Ornithoscelida), with Sauropodomorpha and Herrerasauridae (as the redefined Saurischia) forming its monophyletic outgroup. This new tree topology requires redefinition and rediagnosis of Dinosauria and the subsidiary dinosaurian clades. In addition, it forces re-evaluations of early dinosaur cladogenesis and character evolution, suggests that hypercarnivory was acquired independently in herrerasaurids and theropods, and offers an explanation for many of the anatomical features previously regarded as notable convergences between theropods and early ornithischians.

Effect of cell seeding density on matrix-forming capacity of meniscus fibrochondrocytes and nasal chondrocytes in meniscus tissue engineering.

The presence of intact menisci is imperative for the proper function of the knee joint. Meniscus injuries are often treated by the surgical removal of the damaged tissue, which increases the likelihood of post-traumatic osteoarthritis. Tissue engineering holds great promise in producing viable engineered meniscal tissue for implantation using the patient's own cells; however, the cell source for producing the engineered tissue is unclear. Nasal chondrocytes (NC) possess many attractive features for engineering meniscus. However, in order to validate the use of NC for engineering meniscus fibrocartilage, a thorough comparison of NC and meniscus fibrochondrocytes (MFC) must be considered. Our study presents an analysis of the relative features of NC and MFC and their respective chondrogenic potential in a pellet culture model. We showed considerable differences in the cartilage tissue formed by the two different cell types. Our data showed that NC were more proliferative in culture, deposited more extracellular matrix, and showed higher expression of chondrogenic genes than MFC. Overall, our data suggest that NC produce superior cartilage tissue to MFC in a pellet culture model. In addition, NCs produce higher quality cartilage tissue at higher cell seeding densities during cell expansion.

Does Coronal Plane Malalignment of the Tibial Insert in Total Ankle Arthroplasty Alter Distal Foot Bone Mechanics? A Cadaveric Gait Study.

BACKGROUND: Total ankle arthroplasty (TAA) is becoming a more prevalent treatment for end-stage ankle arthritis. However, the effects of malalignment on TAA remain poorly understood. QUESTIONS/PURPOSES: The purpose of this study was to quantify the mechanical effects of coronal plane malalignment of the tibial insert in TAA using cadaveric gait simulation. Specifically, we asked, is there a change in (1) ankle joint congruency, (2) kinematic joint position, (3) kinematic ROM, (4) peak plantar pressure, and (5) center of pressure with varus and valgus malalignment? METHODS: A modified TAA was implanted into seven cadaveric foot specimens. Wedges were used to simulate coronal plane malalignment of the tibial insert. The degree of malalignment (tibial insert angle [TIA] and talar component angle [TCA]) was quantified radiographically for neutral and 5°, 10°, and 15° varus and valgus wedges. Dynamic walking at 1/6 of physiological speed was simulated using a robotic gait simulator. A motion capture system was used to measure foot kinematics, and a pressure mat was used to measure plantar pressure. Joint congruency was quantified as the difference between TIA and TCA. Continuous joint position, joint ROM, peak plantar pressure, and center of pressure for varus and valgus malalignment compared with neutral alignment were estimated using linear mixed effects regression. Pairwise comparisons between malalignment conditions and neutral were considered significant if both the omnibus test for the overall association between outcome and malalignment and the individual pairwise comparison (adjusted for multiple comparisons within a given outcome) had p ≤ 0.05. RESULTS: Descriptively, the TIA and TCA were both less pronounced than the wedge angle and component incongruence was seen (R = 0.65; p < 0.001). Varus malalignment of the tibial insert shifted the tibiotalar joint into varus and internally rotated the joint. The tibiotalar joint's ROM slightly increased as the TIA shifted into varus (1.3 ± 0.7° [mean ± SD] [95% confidence interval -0.7 to 3.4]; p = 0.03), and the first metatarsophalangeal joint's ROM decreased as the TIA shifted into varus (-1.9 ± 0.9° [95% CI -5.6 to 1.7]; p = 0.007). In the sagittal plane, the naviculocuneiform joint's ROM slightly decreased as the TIA shifted into varus (-0.9 ± 0.4° [95% CI -2.1 to 0.3]; p = 0.017). Hallux pressure increased as the TIA became more valgus (59 ± 50 kPa [95% CI -88 to 207]; p = 0.006). The peak plantar pressure slightly decreased in the third and fourth metatarsals as the TIA shifted into valgus (-15 ± 17° [95% CI -65 to 37]; p = 0.03 and -8 ± 4° [95% CI -17 to 1]; p = 0.048, respectively). The fifth metatarsal's pressure slightly decreased as the TIA shifted into valgus (-18 ± 12 kPa [95% CI -51 to 15]) or varus (-7 ± 18 kPa [95% CI -58 to 45]; p = 0.002). All comparisons were made to the neutral condition. CONCLUSIONS: In this cadaver study, coronal plane malalignment in TAA altered foot kinematics and plantar pressure. In general, varus TAA malalignment led to varus shift and internal rotation of the tibiotalar joint, a slight increase in the tibiotalar ROM, and a slight decrease in the first metatarsophalangeal ROM, while a valgus TAA malalignment was manifested primarily through increased hallux pressure with a slight off-loading of the third and fourth metatarsals. CLINICAL RELEVANCE: This study may increase our understanding of the biomechanical processes that underlie the unfavorable clinical outcomes (such as, poor patient-reported outcomes or implant loosening) that have been associated with coronal plane malalignment of the tibial component in TAA.

Peroxisome Protein Prediction in Drosophila melanogaster.

As a laboratory animal, Drosophila melanogaster has made extensive contributions to understanding many areas of fundamental biology as well as being an effective model for human disease. Until recently, there was relatively little known about fly peroxisomes. There were early studies that examined the role of peroxisome enzymes during development of organs like the eye. However, with the advent of a well-annotated, sequenced genome, several groups have collectively determined, first by sequence homology and increasingly by functional studies, Drosophila Peroxins and related peroxisome proteins. Notably, it was shown that Drosophila peroxisome biogenesis is mediated via a well-conserved PTS1 import system. Although the fly genome encodes a Pex7 homologue, a canonical PTS2 import system does not seem to be conserved in Drosophila. Given the homology between Drosophila and Saccharomyces cerevisiae or Homo sapiens peroxisome biogenesis and function, Drosophila has emerged as an effective multicellular system to model human Peroxisome Biogenesis Disorders. Finally, Drosophila peroxisome research has recently come into its own, facilitating new discoveries into the role of peroxisomes within specific tissues, such as testes or immune cells.

A Systematic Cell-Based Analysis of Localization of Predicted Drosophila Peroxisomal Proteins.

Peroxisomes are membrane-bound organelles found in almost all eukaryotic cells. They perform specialized biochemical functions that vary with organism, tissue or cell type. Mutations in human genes required for the assembly of peroxisomes result in a spectrum of diseases called the peroxisome biogenesis disorders. A previous sequence-based comparison of the predicted proteome of Drosophila melanogaster (the fruit fly) to human proteins identified 82 potential homologues of proteins involved in peroxisomal biogenesis, homeostasis or metabolism. However, the subcellular localization of these proteins relative to the peroxisome was not determined. Accordingly, we tested systematically the localization and selected functions of epitope-tagged proteins in Drosophila Schneider 2 cells to determine the subcellular localization of 82 potential Drosophila peroxisomal protein homologues. Excluding the Pex proteins, 34 proteins localized primarily to the peroxisome, 8 showed dual localization to the peroxisome and other structures, and 26 localized exclusively to organelles other than the peroxisome. Drosophila is a well-developed laboratory animal often used for discovery of gene pathways, including those linked to human disease. Our work establishes a basic understanding of peroxisome protein localization in Drosophila. This will facilitate use of Drosophila as a genetically tractable, multicellular model system for studying key aspects of human peroxisome disease.

A dinosaur missing-link? Chilesaurus and the early evolution of ornithischian dinosaurs.

The enigmatic dinosaur taxon Chilesaurus diegosuarezi was originally described as a tetanuran theropod, but this species possesses a highly unusual combination of features that could provide evidence of alternative phylogenetic positions within the clade. In order to test the relationships of Chilesaurus, we added it to a new dataset of early dinosaurs and other dinosauromorphs. Our analyses recover Chilesaurus in a novel position, as the earliest diverging member of Ornithischia, rather than a tetanuran theropod. The basal position of Chilesaurus within the clade and its suite of anatomical characters suggest that it might represent a 'transitional' taxon, bridging the morphological gap between Theropoda and Ornithischia, thereby offering potential insights into the earliest stages of ornithischian evolution, which were previously obscure. For example, our results suggest that pubic retroversion occurred prior to some of the craniodental and postcranial modifications that previously diagnosed the clade (e.g. the presence of a predentary bone and ossified tendons).

Investigations to extend viability of a rainbow trout primary gill cell culture.

The primary culture of fish gill cells can provide functional, cell diverse, model in vitro platforms able to tolerate an aqueous exposure analogous to in vivo tissues. The utility of such models could be extended to a variety of longer term exposure scenarios if a method could be established to extend culture viability when exposed to water for longer periods. Here we report findings of a series of experiments to establish increased longevity, as monitored by culture transepithelial electrical resistance (TEER) and concurrent histological developments. Experimental cultures improved TEER during apical freshwater exposure for a mean of twelve days, compared to previous viabilities of up to 3 days. Cultures with larger surface areas and the use of trout serum rather than foetal bovine serum (FBS) contributed to the improvement, while perfusion of the intact gill prior to cell harvest resulted in a significantly faster preparation. Detailed scanning electron microscopy analysis of cultures revealed diverse surface structures that changed with culture age. Cultures grown on membranes with an increased porosity, collagen coating or 3D structure were of no benefit compared to standard membranes. Increased culture longevity, achieved in this study and reported for the first time, is a significant breakthrough and opens up a variety of future experimentation that has previously not been possible. The extended viability facilitates exploration of in vitro chronic or pulse-exposure test paradigms, longer term physiological and environmental monitoring studies and the potential for interactive co-culture with other organoid micro-tissues.

Immediate postoperative radiographs after shoulder arthroplasty are often poor quality and do not alter care.

BACKGROUND: It is technically difficult to obtain high-quality, postoperative shoulder radiographs immediately after surgery. Further, poor-quality radiographs may be unlikely to change clinical practice or improve patient outcomes. We therefore questioned the value of routine postoperative radiographs after shoulder arthroplasty. QUESTIONS/PURPOSES: We asked whether (1) postanesthesia care unit (PACU) radiographs can reasonably serve as a baseline for future studies; and (2) routine PACU radiographs change clinical care. (3) We also determined the charges associated with routine PACU radiographs and formal radiographic interpretation of these images. METHODS: We retrospectively compared the radiographs of 283 patients who had shoulder arthroplasties (Group 1) who underwent PACU radiographs with those of 241 patients (Group 2) who had their first postoperative radiographs at a later date. Radiographs were compared for quality, ability to serve as a baseline, and their influence on clinical course. Orthopaedic evaluation of each radiograph and the radiographic report were compared and charges were analyzed. RESULTS: All images in Group 1 were single-view radiographs (88% internal rotation), most were underpenetrated (71%); no images changed postoperative management or were considered adequate to serve as a baseline. Group 2 radiographs were multiview radiographs, and 83% were deemed adequate to serve as baseline radiographs. Radiographic interpretation of immediate postoperative radiographs did not change the clinical course or treatment. The charges billed from radiographic evaluation in this study were $64,524 for Group 1. CONCLUSIONS: Routine PACU radiographs, in the absence of a specific indication, may result in poor-quality images. Elimination of these radiographs and radiographic interpretation after shoulder arthroplasty may reduce charges without changing clinical care. LEVEL OF EVIDENCE: Level III, diagnostic study. See Guidelines for Authors for a complete description of levels of evidence.

Methylphenidate-induced dendritic spine formation and DeltaFosB expression in nucleus accumbens.

Methylphenidate is the psychostimulant medication most commonly prescribed to treat attention deficit hyperactivity disorder (ADHD). Recent trends in the high usage of methylphenidate for both therapeutic and nontherapeutic purposes prompted us to investigate the long-term effects of exposure to the drug on neuronal adaptation. We compared the effects of chronic methylphenidate or cocaine (15 mg/kg, 14 days for both) exposure in mice on dendritic spine morphology and DeltaFosB expression in medium-sized spiny neurons (MSN) from ventral and dorsal striatum. Chronic methylphenidate increased the density of dendritic spines in MSN-D1 (MSN-expressing dopamine D1 receptors) from the core and shell of nucleus accumbens (NAcc) as well as MSN-D2 (MSN-expressing dopamine D2 receptors) from the shell of NAcc. In contrast, cocaine increased the density of spines in both populations of MSN from all regions of striatum. In general, the effect of methylphenidate on the increase of shorter spines (class 2) was less than that of cocaine. Interestingly, the methylphenidate-induced increase in the density of relatively longer spines (class 3) in the shell of NAcc was bigger than that induced by cocaine. Furthermore, methylphenidate exposure increased expression of DeltaFosB only in MSN-D1 from all areas of striatum, and surprisingly, the increase was greater than that induced by cocaine. Thus, our results show differential effects of methylphenidate and cocaine on neuronal adaptation in specific types of MSN in reward-related brain regions.

Towards a more representative in vitro method for fish ecotoxicology: morphological and biochemical characterisation of three-dimensional spheroidal hepatocytes.

The use of fish primary cells and cell lines offer an in vitro alternative for assessment of chemical toxicity and the evaluation of environmental samples in ecotoxicology. However, their uses are not without limitations such as short culture periods and loss of functionality, particularly with primary tissue. While three-dimensional (spheroid) technology is now established for in vitro mammalian toxicity studies, to date it has not been considered for environmental applications in a model aquatic species. In this study we report development of a reproducible six-well plate, gyratory-mediated method for rainbow trout (Oncorhynchus mykiss) hepatocyte spheroid culture and compare their functional and biochemical status with two-dimensional (2D) monolayer hepatocytes. Primary liver spheroid formation was divided into two stages, immature (1-5 days) and mature (≥6 days) according to size, shape and changes in functional and biochemical parameters (protein, glucose, albumin and lactate dehydrogenase). Mature spheroids retained the morphological characteristics (smooth outer surface, tight cell-cell contacts) previously described for mammalian spheroids as demonstrated by light and scanning electron microscopy. Glucose production and albumin synthesis were significantly higher in mature spheroids when compared to conventional 2D monolayer cultures (P < 0.01) and increased as spheroids matured (P < 0.01). Basal lactate dehydrogenase (LDH) leakage significantly decreased during spheroid formation and was significantly lower than 2D cultures (P < 0.01). It is therefore suggested that mature spheroids can maintain a high degree of functional, biochemical and morphological status over-time in culture that is superior to conventional 2D models and can provide realistic organotypic responses in vitro. Trout spheroids that take ~6-8 days to reach maturity would be suitable for use in acute toxicological tests and since it is possible to culture individual spheroids for over a month, there is potential for this work to lead towards in vitro bioaccumulation alternatives and to conduct high throughput screens of chronic exposure. This is an important step forward for developing alternative in vitro tools in future fish ecotoxicological studies.

Intracranial extension of retrobulbar blastomycosis (Blastomyces dermatitidis) in a dog.

Blastomycosis (Blastomyces dermatitidis) is a fungal disease that is endemic in the southern United States. This case report illustrates the clinical, MRI and histopathologic findings in a dog with invasion of a retrobulbar blastomycotic lesion into the calvarium. A 5-year-old intact female Weimaraner was referred for a 2-month history of change in behavior and recent onset of visual deficits. Magnetic resonance imaging (MRI) examination revealed a large (5.8 × 2.0 × 2.5 cm) mass extending from the left orbit through a circular defect in the left cranioventral aspect of the calvarium caudally to the level of the pituitary fossa and interthalamic adhesion. The mass was heterogeneously iso- to hypointense on T2-W images, slightly hypointense on T1-W images, did not attenuate on fluid attenuated inversion recovery (FLAIR) images, and did not show evidence of susceptibility artifact on T2*-W gradient recalled echo (GRE) images. Vasogenic edema and associated mass effect were noted. The mass showed strong homogeneous contrast enhancement with well-defined margins and had thickening of the adjacent meninges (dural tail sign). Based on MRI findings a malignant neoplastic process was considered most likely and the patient was placed on oral prednisone to decrease peri-tumoral inflammation. The dog initially improved but was euthanized 3 weeks later for worsening clinical signs. Histopathologic assessment of the mass revealed marked pyogranulomatous optic neuritis with intralesional fungal yeasts consistent with blastomycosis (Blastomyces dermatitidis). To our knowledge this is the first report of invasion of a retrobulbar blastomycotic lesion into the calvarium in a dog.

Similar patient reported outcomes at long-term follow-up after external fixation versus internal fixation of the anterior ring component of APC injuries.

PURPOSE: Unstable pelvic ring injuries produced by external rotation of the hemipelvis and a symphyseal disruption are most often treated with internal fixation of the anterior ring, with percutaneous treatment of the posterior ring as needed. In some clinical situations, patients are treated with external fixation for their anterior injuries and the long-term functional outcomes associated with external fixation are not well understood. We ask if there is a difference in functional outcome, between treatment of these injuries with internal versus external fixation, when measured at a minimum of three years after injury. METHOD: This was a retrospective cohort study performed at a level one regional trauma center. Trauma database review identified 128 patients, with 70 subsequently excluded, with unstable anterior posterior compression (APC) pelvic ring injuries (OTA 61B2.3 & 61C1.2) treated with surgery with minimum three years of follow-up. An intervention of internal fixation versus external fixation of anterior pelvic ring was performed, and depending on the injury, supplemented with posterior iliosacral screw fixation. Main outcome was measured with the Majeed functional outcome score (0-100). RESULTS: Patients treated with external fixation reported a Majeed score of 70 (95% CI 28-100) compared to 79 (95% CI 36-100) in those with internal fixation (p-value 0.28). Subgroups of the Majeed score were not significantly different (p value > 0.05). Open fractures, severity of injury, and ISS were worse in those treated with external fixation. There was no differential loss to follow-up. Conclusion Patients with unstable pelvic ring injuries with symphyseal disruptions treated with external fixation as definitive treatment versus internal fixation may fare no different in the long term.

Nasal Chondrocyte-Derived Soluble Factors Affect Chondrogenesis of Cocultured Mesenchymal Stem Cells.

To investigate the effect of soluble factors released from human nasal chondrocytes (NCs) on cocultured human bone marrow mesenchymal stem cells (MSCs) and NC tissue-engineered constructs. Cartilage engineered from pure NCs on a three-dimensional (3D) porous collagen scaffold was cultured indirectly in a Transwell system with cartilage engineered from a direct coculture of human bone marrow-derived MSCs and NCs on a 3D porous collagen scaffold. The soluble factors were measured in the conditioned media from the different chambers of the Transwell system. Engineered cartilage from cocultures exposed to the pure NC construct exhibited reduced chondrogenic potential relative to control constructs, shown by reduced extracellular matrix deposition and increased expression of hypertrophic markers. Analysis of the soluble factors within the conditioned media showed an increase in inflammatory cytokines in the coculture chamber exposed to the pure NC construct. Principal component analysis revealed that the majority of the data variance could be explained by proinflammatory factors and hypertrophic chondrogenesis. In conclusion, our data suggest that inflammatory cytokines derived from NCs reduce the chondrogenic potential of coculture engineered cartilage through the induction of hypertrophic chondrogenesis. Impact statement The use of engineered cartilage from cocultured nasal chondrocytes (NCs) and mesenchymal stem cells for nasal cartilage reconstruction may be problematic. Our data suggest that the soluble factors from surrounding native NCs in the cartilage to be fixed can compromise the quality of the engineered cartilage if used in reconstructive surgery.

A neurocomputational method for fully automated 3D dendritic spine detection and segmentation of medium-sized spiny neurons.

Acquisition and quantitative analysis of high resolution images of dendritic spines are challenging tasks but are necessary for the study of animal models of neurological and psychiatric diseases. Currently available methods for automated dendritic spine detection are for the most part customized for 2D image slices, not volumetric 3D images. In this work, a fully automated method is proposed to detect and segment dendritic spines from 3D confocal microscopy images of medium-sized spiny neurons (MSNs). MSNs constitute a major neuronal population in striatum, and abnormalities in their function are associated with several neurological and psychiatric diseases. Such automated detection is critical for the development of new 3D neuronal assays which can be used for the screening of drugs and the studies of their therapeutic effects. The proposed method utilizes a generalized gradient vector flow (GGVF) with a new smoothing constraint and then detects feature points near the central regions of dendrites and spines. Then, the central regions are refined and separated based on eigen-analysis and multiple shape measurements. Finally, the spines are segmented in 3D space using the fast marching algorithm, taking the detected central regions of spines as initial points. The proposed method is compared with three popular existing methods for centerline extraction and also with manual results for dendritic spine detection in 3D space. The experimental results and comparisons show that the proposed method is able to automatically and accurately detect, segment, and quantitate dendritic spines in 3D images of MSNs.

Testing pterosaur ingroup relationships through broader sampling of avemetatarsalian taxa and characters and a range of phylogenetic analysis techniques.

The pterosaurs first appear in the fossil record in the middle of the Late Triassic. Their earliest representatives are known from Northern Hemisphere localities but, by the end of the Jurassic Period, this clade of flying reptiles achieved a global distribution, as well as high levels of diversity and disparity. Our understanding of early pterosaur evolution and the fundamental interrelationships within Pterosauria has improved dramatically in recent decades. However, there is still debate about how the various pterosaur subgroups relate to one another and about which taxa comprise these. Many recent phylogenetic analyses, while sampling well from among the known Triassic and Early Jurassic pterosaurs, have not included many non-pterosaurian ornithodirans or other avemetatarsalians. Given the close relationship between these groups of archosaurs, the omission of other ornithodirans and avemetatarsalians has the potential to adversely affect the results of phylogenetic analyses, in terms of character optimisation and ingroup relationships recovered. This study has addressed this issue and tests the relationships between the early diverging pterosaur taxa following the addition of avemetatarsalian taxa and anatomical characters to an existing early pterosaur dataset. This study has, for the first time, included taxa that represent the aphanosaurs, lagerpetids, silesaurids and dinosaurs, in addition to early pterosaurs. Anatomical characters used in other recent studies of archosaurs and early dinosaurs have also been incorporated. By expanding the outgroup taxa and anatomical character coverage in this pterosaur dataset, better resolution between the taxa within certain early pterosaur subclades has been achieved and stronger support for some existing clades has been found; other purported clades of early pterosaurs have not been found in this analysis-for example there is no support for a monophyletic Eopterosauria or Eudimorphodontidae. Further support has been found for a sister-taxon relationship between Peteinosaurus zambelli and Macronychoptera, a clade here named Zambellisauria (clade nov.), as well as for a monophyletic and early diverging Preondactylia. Some analyses also support the existence of a clade that falls as sister-taxon to the zambellisaurs, here named Caviramidae (clade nov.). Furthermore, some support has been found for a monophyletic Austriadraconidae at the base of Pterosauria. Somewhat surprisingly, Lagerpetidae is recovered outside of Ornithodira sensu stricto, meaning that, based upon current definitions at least, pterosaurs fall within Dinosauromorpha in this analysis. However, fundamental ornithodiran interrelationships were not the focus of this study and this particular result should be treated with caution for now. However, these results do further highlight the need for broader taxon and character sampling in phylogenetic analyses, and the effects of outgroup choice on determining ingroup relationships.

Suppression of Hypertrophy During in vitro Chondrogenesis of Cocultures of Human Mesenchymal Stem Cells and Nasal Chondrocytes Correlates With Lack of in vivo Calcification and Vascular Invasion.

OBJECTIVE: Human nasal septal chondrocytes (NC) are a promising minimally invasive derivable chondrogenic cell source for cartilage repair. However, the quality of NC-derived cartilage is variable between donors. Coculture of NC with mesenchymal stem cells (MSCs) mitigates the variability but with undesirable markers of chondrocyte hypertrophy, such as type X collagen, and the formation of unstable calcifying cartilage at ectopic sites. In contrast, monoculture NC forms non-calcifying stable cartilage. Formation of a stable NC-MSC coculture cartilage is crucial for clinical application. The aim of this study was to explore the utility of parathyroid hormone-related peptide (PTHrP) hormone to suppress chondrocyte hypertrophy in NC-MSC cocultures and form stable non-calcifying cartilage at ectopic sites. METHODS: Human NC and bone marrow MSCs, and cocultures of NC and MSC (1:3 ratio) were aggregated in pellet form and subjected to in vitro chondrogenesis for 3 weeks in chondrogenic medium in the presence and absence of PTHrP. Following in vitro chondrogenesis, the resulting pellets were implanted in immunodeficient athymic nude mice for 3 weeks. RESULTS: Coculture of NC and MSC resulted in synergistic cartilage matrix production. PTHrP suppressed the expression of hypertrophy marker, type X collagen (COL10A1), in a dose-dependent fashion without affecting the synergism in cartilage matrix synthesis, and in vivo calcification was eradicated with PTHrP. In contrast, cocultured control (CC) pellets without PTHrP treatment expressed COL10A1, calcified, and became vascularized in vivo.

The Assessment of Clinically Significant Differences in Treating Spinal Deformity Using the SRS Questionnaire: What Is the Threshold of Change That Is Meaningful to Patients?

BACKGROUND: The measurement of health-related quality of life is important in spinal deformity surgery. The Scoliosis Research Society questionnaire has allowed disease-specific research in this area, and determining the minimal clinically important difference (MCID) is as important as it is elusive. We seek to further refine our estimations of clinically perceived improvements by the patient. METHODS: We used an anchor-based approach for each domain of the SRS questionnaire to compare changes at 1 year after treatment. We set the MCID as the upper 95% boundary of the "no change" group bordering the "improvement" arm, where the patients may start to perceive their own change toward the better. We compared this with the mean change. RESULTS: The threshold value for the MCID was 0.54 for the pain domain, 0.31 for function, 0.62 for self-image, and 0.5 for mental health. The mean changes in our group's pain and self-image exceeded their MCID. CONCLUSIONS: Compared with our previous work, we further attempted to refine our assessment of the MCID in spinal deformity. Pain continues to show clinically significant improvement, and self-image also demonstrated mean improvement over its estimated MCID. LEVEL OF EVIDENCE: 2. CLINICAL RELEVANCE: This result in self-image is an important addition to the MCID literature, given its lack of consistency in previous work.