Clickable Dynamic Bioinks Enable Post-Printing Modifications of Construct Composition and Mechanical Properties Controlled over Time and Space.

Bioprinting is a booming technology, with numerous applications in tissue engineering and regenerative medicine. However, most biomaterials designed for bioprinting depend on the use of sacrificial baths and/or non-physiological stimuli. Printable biomaterials also often lack tunability in terms of their composition and mechanical properties. To address these challenges, the authors introduce a new biomaterial concept that they have termed "clickable dynamic bioinks". These bioinks use dynamic hydrogels that can be printed, as well as chemically modified via click reactions to fine-tune the physical and biochemical properties of printed objects after printing. Specifically, using hyaluronic acid (HA) as a polymer of interest, the authors investigate the use of a boronate ester-based crosslinking reaction to produce dynamic hydrogels that are printable and cytocompatible, allowing for bioprinting. The resulting dynamic bioinks are chemically modified with bioorthogonal click moieties to allow for a variety of post-printing modifications with molecules carrying the complementary click function. As proofs of concept, the authors perform various post-printing modifications, including adjusting polymer composition (e.g., HA, chondroitin sulfate, and gelatin) and stiffness, and promoting cell adhesion via adhesive peptide immobilization (i.e., RGD peptide). The results also demonstrate that these modifications can be controlled over time and space, paving the way for 4D bioprinting applications.

Obesity in adult patients with inflammatory bowel disease: Clinical features and impact on disability. A cross-sectional survey from the GETAID.

BACKGROUND: In recent years, an increasing prevalence of obesity in inflammatory bowel disease (IBD) has been observed. However, only a few studies have focused on the impact of overweight and obesity on IBD-related disability. AIMS: To identify the factors associated with obese and overweight patients with IBD, including IBD-related disability. PATIENTS AND METHODS: In this cross-sectional study, we included 1704 consecutive patients with IBD in 42 centres affiliated with the Groupe d'Etude Therapeutique des Affections Inflammatoires du tube Digestif (GETAID) using a 4-page questionnaire. Factors associated with obesity and overweight were assessed using univariate and multivariate analyses (odds ratios (ORs) are provided with 95% confidence intervals). RESULTS: The prevalence rates of overweight and obesity were 24.1% and 12.2%, respectively. Multivariable analyses were stratified by age, sex, type of IBD, clinical remission and age at diagnosis of IBD. Overweight was significantly associated with male sex (OR = 0.52, 95% CI [0.39-0.68], p < 0.001), age (OR = 1.02, 95% CI [1.01-1.03], p < 0.001) and body image subscore (OR = 1.15, 95% CI [1.10-1.20], p < 0.001) (Table 2). Obesity was significantly associated with age (OR = 1.03, 95% CI [1.02-1.04], p < 0.001), joint pain subscore (OR = 1.08, 95% CI [1.02-1.14], p < 0.001) and body image subscore (OR = 1.25, 95% CI [1.19-1.32], p < 0.001) (Table 3). CONCLUSION: The increasing prevalence of overweight and obesity in patients with IBD is associated with age and poorer body image. A holistic approach to IBD patient care should be encouraged to improve IBD-related disability and to prevent rheumatological and cardiovascular complications.

Determination of the minimal important difference for inspiratory muscle strength in people with severe and very severe COPD.

OBJECTIVE: Inspiratory muscle training is recommended for people with chronic obstructive pulmonary disease (COPD) with inspiratory muscle weakness. Clinical interpretation of changes in inspiratory muscle strength could be helped by the determination of cut-off values. The aim of this study was to estimate the minimal important difference for inspiratory muscle strength assessed with maximal inspiratory pressure (MIP) in people with COPD. DESIGN: Post hoc analysis of a randomized controlled trial (EMI2 study) including people with severe to very severe COPD undergoing a pulmonary rehabilitation program was conducted. The determination of the minimal important difference was realized using both anchor-based and distribution-based methods. SETTING: The study includes patients admitted to the rehabilitation program unit of the Centre Hospitalier des Pays de Morlaix (Morlaix, France) between March 5, 2014 and September 8, 2016. PARTICIPANTS: Seventy-three people with severe to very severe COPD (age 62.2 ± 8.0 years, forced expiratory volume in 1 s 36.4 ± 9.5% of theoretical) were analyzed. INTERVENTION: Patients followed a standardized pulmonary rehabilitation program 5 days a week for 4 weeks. The program included aerobic training, ground-based outdoor walking training, and strengthening of lower and upper limb muscles. MAIN MEASURES: At the end of the pulmonary rehabilitation program, MIP improved by 14.8 ± 14.9 cmH2O (p < 0.05). Regarding the anchor-based method, only the modified Medical Research Council was selected as an appropriate anchor. The receiver operating characteristic curve analysis reported a minimal important difference of 13.5 cmH2O (sensibility: 75% specificity: 67.5%). Using distribution-based methods, the estimate of minimal important difference was 7.9 cmH2O (standard error of measurement method) and 10.9 cmH2O (size effect method). RESULTS: The estimations proposed by this study ranged from 7.9 to 13.5 cmH2O. CONCLUSIONS: The measurement of minimal important difference is a simple tool for assessing the changes of inspiratory muscle strength during a pulmonary rehabilitation program. We propose a minimal important difference of 13.5 cmH2O for the improvement of MIP. Further studies are needed to confirm this estimation.ClinicalTrials.gov identifier: NCT02074813.

Microsatellite Instability and Aberrant Pre-mRNA Splicing: How Intimate Is It?

Cancers that belong to the microsatellite instability (MSI) class can account for up to 15% of all cancers of the digestive tract. These cancers are characterized by inactivation, through the mutation or epigenetic silencing of one or several genes from the DNA MisMatch Repair (MMR) machinery, including MLH1, MLH3, MSH2, MSH3, MSH6, PMS1, PMS2 and Exo1. The unrepaired DNA replication errors turn into mutations at several thousand sites that contain repetitive sequences, mainly mono- or dinucleotides, and some of them are related to Lynch syndrome, a predisposition condition linked to a germline mutation in one of these genes. In addition, some mutations shortening the microsatellite (MS) stretch could occur in the 3'-intronic regions, i.e., in the ATM (ATM serine/threonine kinase), MRE11 (MRE11 homolog) or the HSP110 (Heat shock protein family H) genes. In these three cases, aberrant pre-mRNA splicing was observed, and it was characterized by the occurrence of selective exon skipping in mature mRNAs. Because both the ATM and MRE11 genes, which as act as players in the MNR (MRE11/NBS1 (Nibrin)/RAD50 (RAD50 double strand break repair protein) DNA damage repair system, participate in double strand breaks (DSB) repair, their frequent splicing alterations in MSI cancers lead to impaired activity. This reveals the existence of a functional link between the MMR/DSB repair systems and the pre-mRNA splicing machinery, the diverted function of which is the consequence of mutations in the MS sequences.

LED-pumped Cr:LiSAF laser system operating at 100†Hz based on a multipass amplifier.

The LED-pumping technology is used for the first time, to the best of our knowledge, to develop a complete master oscillator power amplifier (MOPA) system including a multipass amplifier. A pumping head using an original slab architecture is developed integrating a Cr:LiSAF slab pumped by 2112 blue LEDs via a Ce:YAG luminescent concentrator. The slab configuration enables the reaching of a large number of passes-up to 22-together with access to efficient cooling, allowing for a repetition rate scale up. For 22 passes, the amplifier delivers pulses with energy up to 2.4 mJ at 10-Hz repetition rate with a gain of 4.36 at 825 nm. A complete study of the MOPA is described, concluding in nearly constant performances versus the repetition rate, up to 100 Hz.

Design and characterization of an in vivo injectable hydrogel with effervescently generated porosity for regenerative medicine applications.

Injectable hydrogels that polymerize directly in vivo hold significant promises in clinical settings to support the repair of damaged or failing tissues. Existing systems that allow cellular and tissue ingrowth after injection are limited because of deficient porosity and lack of oxygen and nutrient diffusion inside the hydrogels. Here is reported for the first time an in vivo injectable hydrogel in which the porosity does not pre-exist but is formed concomitantly with its in situ injection by a controlled effervescent reaction. The hydrogel tailorable crosslinking, through the reaction of polyethylene glycol with lysine dendrimers, allows the mixing and injection of precursor solutions from a dual-chamber syringe while entrapping effervescently generated CO2 bubbles to form highly interconnected porous networks. The resulting structures allow preserving modular mechanical properties (from 12.7 ± 0.9 to 29.9 ± 1.7 kPa) while being cytocompatible and conducive to swift cellular attachment, proliferation, in-depth infiltration and extracellular matrix deposition. Most importantly, the subcutaneously injected porous hydrogels are biocompatible, undergo tissue remodeling and support extensive neovascularisation, which is of significant advantage for the clinical repair of damaged tissues. Thus, the porosity and injectability of the described effervescent hydrogels, together with their biocompatibility and versatility of mechanical properties, open broad perspectives for various regenerative medicine or material applications, since effervescence could be combined with a variety of other systems of swift crosslinking. STATEMENT OF SIGNIFICANCE: A major challenge in hydrogel design is the synthesis of injectable formulations allowing easy handling and dispensing in the site of interest. However, the lack of adequate porosity inside hydrogels prevent cellular entry and, therefore, vascularization and tissue ingrowth, limiting the regenerative potential of a vast majority of injectable hydrogels. We describe here the development of an acellular hydrogel that can be injected directly in situ while allowing the simultaneous formation of porosity. Such hydrogel would facilitate handling through injection while providing a porous structure supporting vascularization and tissue ingrowth.

Multi-expert annotation of Crohn's disease images of the small bowel for automatic detection using a convolutional recurrent attention neural network.

Background and study aims Computer-aided diagnostic tools using deep neural networks are efficient for detection of lesions in endoscopy but require a huge number of images. The impact of the quality of annotation has not been tested yet. Here we describe a multi-expert annotated dataset of images extracted from capsules from Crohn's disease patients and the impact of the quality of annotations on the accuracy of a recurrent attention neural network. Methods Images of capsule were annotated by a reader first and then reviewed by three experts in inflammatory bowel disease. Concordance analysis between experts was evaluated by Fleiss' kappa and all the discordant images were, again, read by all the endoscopists to obtain a consensus annotation. A recurrent attention neural network developed for the study was tested before and after the consensus annotation. Available neural networks (ResNet and VGGNet) were also tested under the same conditions. Results The final dataset included 3498 images with 2124 non-pathological (60.7 %), 1360 pathological (38.9 %), and 14 (0.4 %) inconclusive. Agreement of the experts was good for distinguishing pathological and non-pathological images with a kappa of 0.79 ( P  < 0.0001). The accuracy of our classifier and the available neural networks increased after the consensus annotation with a precision of 93.7 %, sensitivity of 93 %, and specificity of 95 %. Conclusions The accuracy of the neural network increased with improved annotations, suggesting that the number of images needed for the development of these systems could be diminished using a well-designed dataset.

In Vitro Studies of Xylitol and Erythritol Inhibition of Streptococcus Mutans and Streptococcus Sobrinus Growth and Biofilm Production.

The aim of this study was to evaluate synergy and inhibitory effects of xylitol and erythritol on Streptococcus mutans and Streptococcus sobrinus growth and biomass production on a polystyrene plastic surface. Study design; S. mutans and sobrinus strains (American Type Culture Collection reference strains 31341, 35668, 25175, sobrinus 33478) were cultivated in media (Todd Hewitt Broth with 1% sucrose or heart-brain infusion broth with 1% sucrose) at differing concentrations of xylitol or erythritol in microtiter assay plates incubated for 48 hours. Bacterial growth was quantified and measured by optical density using a microplate reader. Experiments assessing synergy and biofilm growth were carried out also using microdilution assays. All four strains were inhibited by 30% (w/v) xylitol, and 15% erythritol at 150mg/ml erythritol, 2/4 strains had reduced growth; at 270mg/ml, 4/4 strains were inhibited. Bactericidal effects were not observed at any polyol concentration. Combinations of both polyols in a checker board array were used to determine if there were any benefits of polyol combinations. Results The combination studies yielded mixed outcomes with indifference in growth for strains 68 and 78, potential additive effect for strain 75 and possible antagonism for strain 41. Assessment of biomass formation and polyol interference were also performed post MIC assessment. Strains 41, 68 and 75 produced significant biomass in the absence of either polyol. Both polyols inhibited biomass formation in a dose-dependent fashion. Strain 75 is a poor biomass producer and could not be assessed for polyol effects in our assay. Conclusion: Our results demonstrate significant polyol influence on the oral Streptococcal strains tested in our laboratory.

Radiogenomics-based cancer prognosis in colorectal cancer.

Radiogenomics aims at investigating the relationship between imaging radiomic features and gene expression alterations. This study addressed the potential prognostic complementary value of contrast enhanced computed tomography (CE-CT) radiomic features and gene expression data in primary colorectal cancers (CRC). Sixty-four patients underwent CT scans and radiomic features were extracted from the delineated tumor volume. Gene expression analysis of a small set of genes, previously identified as relevant for CRC, was conducted on surgical samples from the same tumors. The relationships between radiomic and gene expression data was assessed using the Kruskal-Wallis test. Multiple testing was not performed, as this was a pilot study. Cox regression was used to identify variables related to overall survival (OS) and progression free survival (PFS). ABCC2 gene expression was correlated with N (p = 0.016) and M stages (p = 0.022). Expression changes of ABCC2, CD166, CDKNV1 and INHBB genes exhibited significant correlations with some radiomic features. OS was associated with Ratio 3D Surface/volume (p = 0.022) and ALDH1A1 expression (p = 0.042), whereas clinical stage (p = 0.004), ABCC2 expression (p = 0.035), and EntropyGLCM_E (p = 0.0031), were prognostic factors for PFS. Combining CE-CT radiomics with gene expression analysis and histopathological examination of primary CRC could provide higher prognostic stratification power, leading to improved patient management.

Laponite nanoparticle-associated silated hydroxypropylmethyl cellulose as an injectable reinforced interpenetrating network hydrogel for cartilage tissue engineering.

Articular cartilage is a connective tissue which does not spontaneously heal. To address this issue, biomaterial-assisted cell therapy has been researched with promising advances. The lack of strong mechanical properties is still a concern despite significant progress in three-dimensional scaffolds. This article's objective was to develop a composite hydrogel using a small amount of nano-reinforcement clay known as laponites. These laponites were capable of self-setting within the gel structure of the silated hydroxypropylmethyl cellulose (Si-HPMC) hydrogel. Laponites (XLG) were mixed with Si-HPMC to prepare composite hydrogels leading to the development of a hybrid interpenetrating network. This interpenetrating network increases the mechanical properties of the hydrogel. The in vitro investigations showed no side effects from the XLG regarding cytocompatibility or oxygen diffusion within the composite after cross-linking. The ability of the hybrid scaffold containing the composite hydrogel and chondrogenic cells to form a cartilaginous tissue in vivo was investigated during a 6-week implantation in subcutaneous pockets of nude mice. Histological analysis of the composite constructs revealed the formation of a cartilage-like tissue with an extracellular matrix containing glycosaminoglycans and collagens. Overall, this new hybrid construct demonstrates an interpenetrating network which enhances the hydrogel mechanical properties without interfering with its cytocompatibility, oxygen diffusion, or the ability of chondrogenic cells to self-organize in the cluster and produce extracellular matrix components. This composite hydrogel may be of relevance for the treatment of cartilage defects in a large animal model of articular cartilage defects. STATEMENT OF SIGNIFICANCE: Articular cartilage is a tissue that fails to heal spontaneously. To address this clinically relevant issue, biomaterial-assisted cell therapy is considered promising but often lacks adequate mechanical properties. Our objective was to develop a composite hydrogel using a small amount of nano reinforcement (laponite) capable of gelling within polysaccharide based self-crosslinking hydrogel. This new hybrid construct demonstrates an interpenetrating network (IPN) which enhances the hydrogel mechanical properties without interfering with its cytocompatibility, O2 diffusion and the ability of chondrogenic cells to self-organize in cluster and produce extracellular matrix components. This composite hydrogel may be of relevance for the treatment of cartilage defects and will now be considered in a large animal model of articular cartilage defects.

Nucleotide and Nucleotide Sugar Analysis in Cell Extracts by Capillary Electrophoresis.

In biotechnological processes the intracellular level of nucleotides and nucleotide sugars have a direct impact on the post-translational modification (glycosylation) of the therapeutic protein products and on the exopolysaccharide pattern of the cells. Thus, they are precursors and also key components in the production of glycoproteins and glycolipids. All four nucleotides (at different phosphorylation stages) and their natural sugar derivatives coexist in biological samples. Their relative ratios depend on the actual conditions under which the cells are grown. Therefore, their simultaneous determination at different time points and different cell culture conditions in biotechnological samples is of interest in order to develop the optimal cell culture process. In our study capillary electrophoresis (CE) combined with UV detection @ 260 nm was selected for the separation and quantification of the complex nucleotide mixture of the structurally very similar nucleotides and nucleotide sugars in cell extracts. The high separation efficiency of CE as well as its insensitivity to the complex cell matrix makes this method superior to commonly used HPLC methods. In our study eleven nucleotides and six nucleotide sugars were analyzed. A robust and reproducible analysis system was developed. As background electrolyte borate (40 mM, pH 9.5) was used containing 1% PEG (MW 35'000 Da) which enhanced resolution. In order to obtain high reproducibility in terms of migration time, mandatory for the unambiguous identification of the single compounds in the complex cell extract mixtures, dynamic coating was also employed. The method was tested for CHO cell extracts where three sugar nucleotides and seven nucleotides were identified and quantified using GDP-Glc as internal standard.

Water-soluble nickel-bis(dithiolene) complexes as photothermal agents.

The remarkable photothermal properties of pegylated nickel-bis(dithiolene) complexes in water can induce cell death under laser irradiation in the near infrared (NIR) region.

Myogenic and nonmyogenic cells differentially express proteinases, Hsc/Hsp70, and BAG-1 during skeletal muscle regeneration.

Skeletal muscle has a remarkable capacity to regenerate after injury. To determine whether changes in the expression of proteinases, 73-kDa constitutive heat shock cognate protein (Hsc70) and stress-inducible 72-kDa heat shock protein (Hsp70) (Hsc/Hsp70), and Bcl-2-associated gene product-1 (BAG-1) contribute to the remodeling response of muscle tissue, tibialis anterior muscles of male Sprague-Dawley rats were injected with 0.75% bupivacaine and removed at 3, 5, 7, 10, 14, 21, or 35 days postinjection (n = 5-7/group). The immunohistochemical analysis of desmin, alpha-actin, and developmental/neonatal myosin heavy chain expressions indicated the presence of myoblasts (days 3-7), inflammatory cells (days 3-7), degenerating myofibers (days 3-7), regenerating myofibers (days 5-10), and growing mature myofibers (days 10-21) in regenerating muscles. Our biochemical analysis documented profound adaptations in proteolytic metabolism characterized by significant increases in the enzyme activities of matrix metalloproteinases 2 and 9 and plasminogen activators (days 3-14), calpains 1 and 2 (days 3-7), cathepsins B and L(days 3-10), and proteasome (days 3-14). Proteasome activity was strongly correlated with proliferating cell nuclear antigen protein level, suggesting that proteasome played a key role in myoblast proliferation. The expression pattern of BAG-1, a regulatory cofactor of Hsc/Hsp70 at the interface between protein folding and proteasomal proteolysis, did not corroborate the changes in proteasome enzyme activity, suggesting that BAG-1 may promote other functions, such as the folding capacity of Hsc/Hsp70. Altogether, the diversity of functions attributed to proteinases in the present study was strongly supported by the relative changes in the proportion of myogenic and nonmyogenic cells over the time course of regeneration.