Dependence of Cartilage Matrix Composition on Biosynthesis,Diffusion, and Reaction

The steady-state composition of articular cartilage has previously been described as a balance between the metabolism of matrix molecules and the loss of these components from the tissue. Single compartment kinetic models have previously been developed to describe the relationship between these processes and overall (spatially-averaged) concentration of matrix molecules. Here, we develop a continuum model to describe the relationship between spatially-varying matrix concentrations and the processes of matrix formation, binding, degradation, and molecular transport within and from the cartilage tissue. At steady-state, the resultant concentration profile, and also spatially-averaged concentration, are predicted to depend on the balance between diffusivity and binding rate, diffusivity and formation rate, and various rate processes, some of which depend on tissue thickness. The predicted concentration profile, for certain parameter values, exhibits similarities to that known to exist for the proteoglycan matrix component, suggesting that transport factors may play an important role in causing the spatial variation in this component. Under other conditions, the predicted concentration profile may have a large portion of bound components and be relatively constant, consistent with the known distribution of collagen in cartilage. Thus, the continuum model may provide insight into the biophysical mechanism underlying matrix distribution within different regions of articular cartilage.     

New Promise and Opportunities for Allosteric Kinase Inhibitors

Drugs that function through allosteric inhibition of kinase signaling represent a promising approach for the targeted discovery of therapeutics. The majority of developed allosteric kinase inhibitors are characterized as type III and IV inhibitors that show good kinome selectivity but generally lack the subtype selectivity of same kinase family. Recently allosteric inhibitors have been developed that bind outside the catalytic kinase domain with high selectivity for specific kinase subtypes. Allosteric inhibitors that bind to the pseudokinase domain of pseudokinase or the extracellular domain of receptor tyrosine kinases are reviewed. We also review recent developments in the field of allosteric kinase inhibitors including examples of proteolysis targeting chimeras, and highlight the unique binding modes for each type of inhibitors and address future opportunities in this area.     

Transformation of Receptor Tyrosine Kinases into Glutamate Receptors and Photoreceptors

Receptor tyrosine kinases (RTKs) are key regulators of cellular functions in metazoans. In vertebrates, RTKs are mostly activated by polypeptides but are not naturally sensitive to amino acids or light. Taking inspiration from Venus kinase receptors (VKRs), an atypical family of RTKs found in nature, we have transformed the human insulin (hIR) and hepatocyte growth factor receptor (hMET) into glutamate receptors by replacing their extracellular binding domains with the ligand‐binding domain of metabotropic glutamate receptor type 2 (mGluR2). We then imparted light sensitivity through covalent attachment of a synthetic glutamate‐based photoswitch via a self‐labelling SNAP tag. By employing a Xenopus laevis oocyte kinase activity assay, we demonstrate how these chimeric RTKs, termed light‐controlled human insulin receptor (LihIR) and light‐controlled human MET receptor (LihMET), can be used to exert optical control over the insulin or MET signaling pathways. Our results outline a potentially general strategy to convert RTKs into photoreceptors.     

Fibrin Matrices as (Injectable) Biomaterials: Formation,Clinical Use,and Molecular Engineering

This review focuses on fibrin, starting from biological mechanisms (its production from fibrinogen and its enzymatic degradation), through its use as a medical device and as a biomaterial, and finally discussing the techniques used to add biological functions and/or improve its mechanical performance through its molecular engineering. Fibrin is a material of biological (human, and even patient's own) origin, injectable, adhesive, and remodellable by cells; further, it is nature's most common choice for an in situ forming, provisional matrix. Its widespread use in the clinic and in research is therefore completely unsurprising. There are, however, areas where its biomedical performance can be improved, namely achieving a better control over mechanical properties (and possibly higher modulus), slowing down degradation or incorporating cell‐instructive functions (e.g., controlled delivery of growth factors). The authors here specifically review the efforts made in the last 20 years to achieve these aims via biomimetic reactions or self‐assembly, as much via formation of hybrid materials.     

Synthesis of Densely Phosphorylated Bis‐1,5‐Diphospho‐myo‐Inositol Tetrakisphosphate and its Enantiomer by Bidirectional P‐Anhydride Formation

The ubiquitous mammalian signaling molecule bis‐diphosphoinositol tetrakisphosphate (1,5‐(PP)₂‐myo‐InsP₄, or InsP₈) displays the most congested three‐dimensional array of phosphate groups found in nature. The high charge density, the accumulation of unstable P‐anhydrides and P‐esters, the lack of UV absorbance, and low levels of optical rotation constitute severe obstacles to its synthesis, characterization, and purification. Herein, we describe the first procedure for the synthesis of enantiopure 1,5‐(PP)₂‐myo‐InsP₄ and 3,5‐(PP)₂‐myo‐InsP₄ utilizing a C₂‐symmetric P‐amidite for desymmetrization and concomitant phosphitylation followed by a one‐pot bidirectional P‐anhydride‐forming reaction that combines sixteen chemical transformations with high efficiency. The configuration of these materials is unambiguously shown by subsequent X‐ray analyses of both enantiomers after being individually soaked into crystals of the kinase domain of human diphosphoinositol pentakisphosphate kinase 2.     

A Multinuclear Metal Complex Based DNase‐Mimetic Artificial Enzyme: Matrix Cleavage for Combating Bacterial Biofilms

Extracellular DNA (eDNA) is an essential structural component during biofilm formation, including initial bacterial adhesion, subsequent development, and final maturation. Herein, the construction of a DNase‐mimetic artificial enzyme (DMAE) for anti‐biofilm applications is described. By confining passivated gold nanoparticles with multiple cerium(IV) complexes on the surface of colloidal magnetic Fe₃O₄ /SiO₂ core/shell particles, a robust and recoverable artificial enzyme with DNase‐like activity was obtained, which exhibited high cleavage ability towards both model substrates and eDNA. Compared to the high environmental sensitivity of natural DNase in anti‐biofilm applications, DMAE exhibited a much better operational stability and easier recoverability. When DMAE was coated on substratum surfaces, biofilm formation was inhibited for prolonged periods of time, and the DMAE excelled in the dispersion of established biofilms of various ages. Finally, the presence of DMAE remarkably potentiated the efficiency of traditional antibiotics to kill biofilm‐encased bacteria and eradiate biofilms.     

Cytosolic accumulation of gammaH2AX is associated with tropomyosin-related kinase A-induced cell death in U2OS cells

Tropomyosin-related kinase A (TrkA) plays an important role in cell survival, differentiation, and apoptosis in various neuronal and nonneuronal cell types. Here we show that TrkA overexpression by the Tet-On system mimics NGF-mediated activation pathways in the absence of nerve growth factor (NGF) stimulation in U2OS cells. In addition, p53 upregulation upon DNA damage was inhibited by TrkA, and p21 was upregulated by TrkA in a p53-independent manner. TrkA overexpression caused cell death by interrupting cell cycle progression, and TrkA-induced cell death was diminished in the presence of its specific inhibitor GW441756. Interestingly, TrkA-mediated cell death was strongly related to gammaH2AX production and poly (ADP-ribose) polymerase cleavage in the absence of DNA damage inducer. In this study, we also reveal that gammaH2AX production by TrkA is blocked by TrkA kinase inhibitors K-252a and GW441756, and it is also significantly inhibited by JNK inhibitor SP600125. Moreover, reduction of cell viability by TrkA was strongly suppressed by SP600125 treatment, suggesting a critical role of JNK in TrkA-induced cell death. We also found that gammaH2AX and TrkA were colocalized in cytosol in the absence of DNA damage, and the nuclear localization of gammaH2AX induced by DNA damage was partly altered to cytosol by TrkA overexpression. Our results suggest that the abnormal cytosolic accumulation of gammaH2AX is implicated in TrkA-induced cell death in the absence of DNA damage.     

The asymptotic variance rate of the output process of finite capacity birth-death queues

We analyze the output process of finite capacity birth-death Markovian queues. We develop a formula for the asymptotic variance rate of the form λ ^*+∑v _i where λ ^* is the rate of outputs and v _i are functions of the birth and death rates. We show that if the birth rates are non-increasing and the death rates are non-decreasing (as is common in many queueing systems) then the values of v _i are strictly negative and thus the limiting index of dispersion of counts of the output process is less than unity. In the M/M/1/K case, our formula evaluates to a closed form expression that shows the following phenomenon: When the system is balanced, i.e. the arrival and service rates are equal, is minimal. The situation is similar for the M/M/c/K queue, the Erlang loss system and some PH/PH/1/K queues: In all these systems there is a pronounced decrease in the asymptotic variance rate when the system parameters are balanced. Research supported in part by Israel Science Foundation Grant 249/02 and 454/05 and by European Network of Excellence Euro-NGI.