Novel In Vitro Platform for Studying the Cell Response to Healthy and Diseased Tendon Matrices.

Current in vitro models poorly represent the healthy or diseased tendon microenvironment, limiting the translation of the findings to clinics. The present work aims to establish a physiologically relevant in vitro tendon platform that mimics biophysical aspects of a healthy and tendinopathic tendon matrix using a decellularized bovine tendon and to characterize tendon cells cultured using this platform. Bovine tendons were subjected to various decellularization techniques, with the efficacy of decellularization determined histologically. The biomechanical and architectural properties of the decellularized tendons were characterized using an atomic force microscope. Tendinopathy-mimicking matrices were prepared by treating the decellularized tendons with collagenase for 3 h or collagenase-chondroitinase (CC) for 1 h. The tendon tissue collected from healthy and tendinopathic patients was characterized using an atomic force microscope and compared to that of decellularized matrices. Healthy human tendon-derived cells (hTDCs) from the hamstring tendon were cultured on the decellularized matrices for 24 or 48 h, with cell morphology characterized using f-actin staining and gene expression characterized using real-time PCR. Tendon matrices prepared by freeze-thawing and 48 h nuclease treatment were fully decellularized, and the aligned structure and tendon stiffness (1.46 MPa) were maintained. Collagenase treatment prepared matrices with a disorganized architecture and reduced stiffness (0.75 MPa), mimicking chronic tendinopathy. Treatment with CC prepared matrices with a disorganized architecture without altering stiffness, mimicking early tendinopathy (1.52 MPa). hTDCs on a healthy tendon matrix were elongated, and the scleraxis (SCX) expression was maintained. On tendinopathic matrices, hTDCs had altered morphological characteristics and lower SCX expression. The expression of genes related to actin polymerization, matrix degradation and remodeling, and immune cell invasion were higher in hTDCs on tendinopathic matrices. Overall, the present study developed a physiological in vitro system to mimic healthy tendons and early and late tendinopathy, and it can be used to better understand tendon cell characteristics in healthy and diseased states.

Chondrocyte-specific response to stiffness-mediated primary cilia formation and centriole positioning.

Mechanical stress and the stiffness of the extracellular matrix are key drivers of tissue development and homeostasis. Aberrant mechanosensation is associated with a wide range of pathologies, including osteoarthritis. Matrix (or substrate) stiffness plays a major role in cell spreading, adhesion, proliferation, and differentiation. However, how specific cells sense substrate stiffness still remains unclear. The primary cilium is an essential cellular organelle that senses and integrates mechanical and chemical signals from the extracellular environment. We hypothesized that the primary cilium dynamically alters its length and position to fine-tune cell mechanosignaling based on substrate stiffness alone. We used a hydrogel system of varying substrate stiffness to examine the role of stiffness on cilia frequency, length, and centriole position as well as cell and nuclei area over time. Contrary to other cell types, we show that chondrocyte primary cilia shorten on softer substrates, demonstrating tissue-specific mechanosensing that is aligned with the tissue stiffness the cells originate from. We further show that stiffness determines centriole positioning to either the basal or apical membrane during attachment and spreading, with centrioles positioned toward the basal membrane on stiffer substrates. These phenomena are mediated by force generation actin-myosin stress fibers in a time-dependent manner. Finally, we show on stiff substrates that primary cilia are involved in tension-mediated cell spreading. We propose that substrate stiffness plays a role in cilia positioning, regulating cellular responses to external forces, and maybe a key driver of mechanosignaling-associated diseases.

Changes in Physiological Tendon Substrate Stiffness Have Moderate Effects on Tendon-Derived Cell Growth and Immune Cell Activation.

Tendinopathy is characterised by pathological changes in tendon matrix composition, architecture, and stiffness, alterations in tendon resident cell characteristics, and fibrosis, with inflammation also emerging as an important factor in tendinopathy progression. The sequence of pathological changes in tendinopathy and the cellular effects of the deteriorating matrix are largely unknown. This study investigated the effects of substrate stiffness on tendon-derived cells (TDCs) and THP-1 macrophages using PDMS substrates representing physiological tendon stiffness (1.88 MPa), a stiff gel (3.17 MPa) and a soft gel (0.61 MPa). Human TDCs were cultured on the different gel substrates and on tissue culture plastic. Cell growth was determined by alamarBlue™ assay, cell morphology was analysed in f-actin labelled cells, and phenotypic markers were analysed by real-time PCR. We found that in comparison to TDCs growing on gels with physiological stiffness, cell growth increased on soft gels at 48 h (23%, p = 0.003). Cell morphology was similar on all three gels. SCX expression was slightly reduced on the soft gels (1.4-fold lower, p = 0.026) and COL1A1 expression increased on the stiff gels (2.2-fold, p = 0.041). Culturing THP-1 macrophages on soft gels induced increased expression of IL1B (2-fold, p = 0.018), and IL8 expression was inhibited on the stiffer gels (1.9-fold, p = 0.012). We also found that culturing TDCs on plastic increased cell growth, altered cell morphology, and inhibited the expression of SCX, SOX9, MMP3, and COL3. We conclude that TDCs and macrophages respond to changes in matrix stiffness. The magnitude of responses measured in TDCs were minor on the range of substrate stiffness tested by the gels. Changes in THP-1 macrophages suggested a more inflammatory phenotype on substrates with non-physiological stiffness. Although cell response to subtle variations in matrix stiffness was moderate, it is possible that these alterations may contribute to the onset and progression of tendinopathy.

A Global Trend Analysis of Kidney Cancer Incidence and Mortality and Their Associations with Smoking, Alcohol Consumption, and Metabolic Syndrome.

BACKGROUND: Kidney cancer is a major urological disease globally, with more than 400 000 new cases diagnosed every year. OBJECTIVE: To investigate incidence and mortality trends for kidney cancer and their associations with modifiable risk factors for kidney cancer. DESIGN, SETTING, AND PARTICIPANTS: The most up-to-date figures on kidney cancer incidence and mortality were collected from the GLOBOCAN database and the Cancer Incidence in Five Continents (CI5). Data on total alcohol consumption and the prevalence of smoking, overweight, diabetes, and hypertension were extracted from the World Health Organization Global Health Observatory data repository. OUTCOME MEASUREMENTS AND STATISTICAL ANALYSIS: Age-standardized rates (ASRs) for incidence and mortality and their correlations with potential risk factors for kidney cancer were investigated. Multivariable linear regression analysis was also conducted. The 10-yr temporal patterns for incidence are presented as the average annual percent change with 95% confidence interval using joinpoint regression analysis. RESULTS AND LIMITATIONS: Globally, there is wide variation in kidney cancer incidence and mortality. There were positive correlations between rates of smoking, alcohol consumption, and overweight and ASRs of kidney cancer incidence and mortality. Multivariable regression analysis revealed that alcohol consumption and overweight were significant risk factors for kidney cancer incidence, while smoking and alcohol consumption were significant risk factors for kidney cancer mortality. There was an increasing trend for the incidence of kidney cancer globally, with a particularly prominent trend for European countries. Of note, increasing incidence of kidney cancer is evident even for younger individuals aged <50 yr. However, cancer registries vary by country and period and there is a lack of data regarding the severity of risk factors and disease characteristics such as the distribution of histological groups, tumor grading, and staging. CONCLUSIONS: There is an increasing trend for kidney cancer incidence globally, particularly in European countries and the younger population. Modifiable risk factors for kidney cancer incidence and mortality have been identified. The increasing incidence of kidney cancer among younger individuals is worrying and warrants early action on possible preventive measures. PATIENT SUMMARY: The incidence of kidney cancer has been increasing globally, particularly in European countries and the younger population. Risk factors include smoking, alcohol consumption, overweight, and hypertension, and these factors are all modifiable.

A Novel In Vitro and In Silico System for Analyzing Complex Mechanobiological Behavior of Chondrocytes in Three-Dimensional Hydrogel Constructs.

Physiological loading is essential for the maintenance of articular cartilage through the regulation of tissue remodeling. To correctly understand the behavior of chondrocytes in their native environment, cell stimulating devices and bioreactors have been developed to examine the effect of mechanical stimuli on chondrocytes. This study describes the design and validation of a novel system for analyzing chondrocyte deformation patterns. This involves an in vitro mechanical device for a controlled application of multi-axial-loading regimes to chondrocyte-seeded agarose constructs and in silico models for analyzing chondrocyte deformation patterns. The computer-controlled device precisely applies compressive, tensile, and shear strains to hydrogel constructs using a customizable macro-based program. The synchronization of the displacements is shown to be accurate with a 1.2% error and is highly reproducible. The device design allows housing for up to eight novel designed free-swelling three-dimensional hydrogel constructs. Constructs include mesh ends and are optimized to withstand the application of up to 7% mechanical tensile and 15% shear strains. Constructs were characterized through mapping the strain within as mechanical load was applied and was validated using light microscopy methods, chondrocyte viability using live/dead imaging, and cell deformation strains. Images were then analyzed to determine the complex deformation strain patterns of chondrocytes under a range of dynamic mechanical stimulations. This is one of the first systems that have characterized construct strains to cellular strains. The features in this device make the system ideally suited for a systematic approach for the investigation of the response of chondrocytes to a complex physiologically relevant deformation profile.

Does theory of planned behaviour play a role in predicting uptake of colorectal cancer screening? A cross-sectional study in Hong Kong.

OBJECTIVE: Colorectal cancer (CRC) ranked second in terms of cancer mortality worldwide. It is associated with a substantial global disease burden. We aimed to examine whether the theory of planned behaviour (TPB) could predict the uptake of faecal immunochemical test to inform novel strategies for enhancing CRC screening participation in population-based programmes. DESIGN: Cross-sectional study. SETTINGS: A Hong Kong-based and territory-wide telephone survey was conducted during the study period from October 2017 to November 2018. PARTICIPANTS: 4800 asymptomatic individuals aged 61-70 years who can communicate in Cantonese were recruited during the survey period. Those who had a history of CRC, chronic bowel inflammation, two or more first-degree relatives with CRC, and received colonoscopy in the past 10 years or faecal occult blood test in the past 5 years were excluded. OUTCOME MEASURES: The association between CRC screening uptake and the factors pertinent to TPB was analysed by univariable and multivariable regression models and the mediating effect of intention. We adjusted for age, gender, educational level, marital and working status, as well as household income. RESULTS: Multivariable regression analysis showed that high perceived behavioural control (adjusted OR (AOR)=12.35, 95% CI 8.21 to 18.60, p<0.001), high intention for CRC screening (AOR=7.86, 95% CI 6.60 to 9.36, p<0.001) and positive attitude towards CRC screening (accuracy and effectiveness: AOR=1.19, 95% CI 1.03 to 1.38, p<0.05; embarrassment and apprehension: AOR=4.27, 95% CI 3.13 to 5.82, p<0.001) were significantly associated with CRC screening uptake. Mediation analysis found that the effect of social norms on screening behaviour was primarily mediated by intention (83.2%), and this indirect, mediated effect accounted for 21.7% to 24.1% of total effects of other constructs in TPB on screening behaviour. CONCLUSIONS: The variables pertinent to TPB could successfully predict CRC screening uptake. Promotion of CRC screening based on interventions that increase perceived behavioural control and behavioural intention could potentially enhance screening uptake. Further studies are needed to establish the cause and effect relationship among these variables and screening uptake, as well as to evaluate the cost-effectiveness of such interventions.

The impact of light during the night.

Exposing chicks to one hour of light during the night disrupts the release of a hormone that is needed by cells in the developing brain to survive.

A novel histone H4 variant H4G regulates rDNA transcription in breast cancer.

Histone variants, present in various cell types and tissues, are known to exhibit different functions. For example, histone H3.3 and H2A.Z are both involved in gene expression regulation, whereas H2A.X is a specific variant that responds to DNA double-strand breaks. In this study, we characterized H4G, a novel hominidae-specific histone H4 variant. We found that H4G is expressed in a variety of human cell lines and exhibit tumor-stage dependent overexpression in tissues from breast cancer patients. We found that H4G localized primarily to the nucleoli of the cell nucleus. This localization was controlled by the interaction of the alpha-helix 3 of the histone fold motif with a histone chaperone, nucleophosmin 1. In addition, we found that modulating H4G expression affects rRNA expression levels, protein synthesis rates and cell-cycle progression. Our data suggest that H4G expression alters nucleolar chromatin in a way that enhances rDNA transcription in breast cancer tissues.

Does the Modified Gartland Classification Clarify Decision Making?

HYPOTHESIS: The modified Gartland classification system for pediatric supracondylar fractures is often utilized as a communication tool to aid in determining whether or not a fracture warrants operative intervention. This study sought to determine the interobserver and intraobserver reliability of the Gartland classification system, as well as to determine whether there was agreement that a fracture warranted operative intervention regardless of the classification system. METHODS: A total of 200 anteroposterior and lateral radiographs of pediatric supracondylar humerus fractures were retrospectively reviewed by 3 fellowship-trained pediatric orthopaedic surgeons and 2 orthopaedic residents and then classified as type I, IIa, IIb, or III. The surgeons then recorded whether they would treat the fracture nonoperatively or operatively. The κ coefficients were calculated to determine interobserver and intraobserver reliability. RESULTS: Overall, the Wilkins-modified Gartland classification has low-moderate interobserver reliability (κ=0.475) and high intraobserver reliability (κ=0.777). A low interobserver reliability was found when differentiating between type IIa and IIb (κ=0.240) among attendings. There was moderate-high interobserver reliability for the decision to operate (κ=0.691) and high intraobserver reliability (κ=0.760). Decreased interobserver reliability was present for decision to operate among residents. For fractures classified as type I, the decision to operate was made 3% of the time and 27% for type IIa. The decision was made to operate 99% of the time for type IIb and 100% for type III. SUMMARY: There is almost full agreement for the nonoperative treatment of Type I fractures and operative treatment for type III fractures. There is agreement that type IIb fractures should be treated operatively and that the majority of type IIa fractures should be treated nonoperatively. However, the interobserver reliability for differentiating between type IIa and IIb fractures is low. Our results validate the Gartland classfication system as a method to help direct treatment of pediatric supracondylar humerus fractures, although the modification of the system, IIa versus IIb, seems to have limited reliability and utility. Terminology based on decision to treat may lead to a more clinically useful classification system in the evaluation and treatment of pediatric supracondylar humerus fractures. LEVEL OF EVIDENCE: Level III-diagnostic studies.

Surgical Anatomy of the Supraclavicular Brachial Plexus.

BACKGROUND: Brachial plexus exploration is performed in infants when addressing birth palsies and in children and adults following trauma. The upper trunk is most often injured. Traditional drawings of the brachial plexus depict the suprascapular nerve as a branch of the midportion of the upper trunk, with the more lateral branch of the upper trunk as the anterior division. We have not found this orientation to be accurate in clinical practice. The purpose of this study was to determine the branching patterns of the upper trunk and to delineate nerve orientations at the level of the divisions. METHODS: Bilateral brachial plexus dissections were performed on eight adult cadavers. The length of the upper trunk and distance of the takeoff of the suprascapular nerve from the anterior and posterior divisions was measured. The native positions of the divisions and of the suprascapular nerve from lateral to medial were recorded across all trunks. RESULTS: In six (38%) of the sixteen specimens, a trifurcation was found at the level of the upper trunk. The suprascapular nerve was the most lateral structure at the clavicular level in all specimens, followed by the posterior division and then the anterior division. The mean distance of the takeoff of the suprascapular nerve was 4 mm proximal to the branch point of the divisions; however, in two specimens, the nerve was found to take off from the posterior division proper. CONCLUSIONS: These findings differ from the standard illustrations and descriptions of the brachial plexus. A thorough understanding of the course of the upper trunk and its branches, such as the suprascapular nerve, is vital to performing nerve transfer surgery or neuroma excision and grafting.

Selection and use of honey as an antioxidant in a French salad dressing system.

Honeys from various floral sources were analyzed to select for utilization as a sweetener and potential source of antioxidants in the formulation of a salad dressing. On the basis of various indicators of potential antioxidant effectiveness, such as the ORAC (oxygen radical absorbance capacity) assay and identification of phenolic profile carried out by HPLC analysis, clover and blueberry honeys were selected. Dressings were stored under accelerated conditions (37 degrees C) for six weeks and at ambient (23 degrees C) and refrigeration (4 degrees C) temperatures for one year. Salad dressings incorporating honey provided protection against oxidation to a degree similar to that of EDTA, as determined by peroxide value and p-anisidine value. This demonstrates the potential for honey to be used as a substitute for EDTA and sweetener (such as HFCS) in commercial salad dressings.

High prevalence and correlates of low bone mineral density in young adults with sickle cell disease.

Sickle cell disease (SCD) is a prevalent genetic disorder in which sickle hemoglobin leads to tissue hypoxia and adverse effects on bone. Published studies suggest that children with SCD often have undiagnosed osteopenia or osteoporosis. Minimal data exist on the prevalence of low bone mineral density (BMD) in adults. Our objective was to describe the prevalence of osteopenia and osteoporosis in adults with SCD and to identify patient or disease characteristics associated with low BMD. We conducted a cross-sectional study of adults with SCD. Through questionnaires, we collected data about disease course and osteoporosis risk factors. Patients underwent dual X-ray absorptiometry (DXA) measurement of BMD at the hip, spine, and forearm and sampling of blood and urine for markers of bone turnover, sickle cell disease severity, and secondary causes of osteoporosis. Our main outcome measure was prevalence of osteopenia and osteoporosis as defined by WHO criteria. Of 32 adults with SCD (14 men and 18 women) with a mean age of 34 years, 72% (95% confidence interval 53-86%) had low BMD at one or more anatomic sites. Thirteen patients were classified as osteoporotic and 10 as osteopenic. The prevalence of low BMD was greatest in the lumbar spine (66% of patients). Significant correlates of decreased BMD included low BMI (P < 0.01), male sex (P = 0.02), and low serum zinc concentrations (P < 0.01). The prevalence of osteopenia and osteoporosis in young adults with SCD is extremely high. Further research is needed to address fracture risk and therapeutic interventions.

Bisphosphonate-associated scleritis: a case report and review.

An 86-year-old female was treated for osteoporosis with alendronate, an aminobisphosphonate. Six weeks after alendronate therapy began, scleritis developed in the patient's right eye. The alendronate was discontinued, and, with the administration of prednisone, the symptoms of scleritis fully resolved. Rechallenge resulted in recurrence of the symptoms. Bisphosphonates are commonly used in the management of osteoporosis and are generally safe agents. Rarely, they have been linked with ocular inflammation. This case report reviews the literature on the ocular effects of bisphosphonates and discusses a possible mechanism for the association.