Endogenous tissue engineering: PTH therapy for skeletal repair

Based on its proven anabolic effects on bone in osteoporosis patients, recombinant parathyroid hormone (PTH_1-34) has been evaluated as a potential therapy for skeletal repair. In animals, the effect of PTH_1-34 has been investigated in various skeletal repair models such as fractures, allografting, spinal arthrodesis and distraction osteogenesis. These studies have demonstrated that intermittent PTH_1-34 treatment enhances and accelerates the skeletal repair process via a number of mechanisms, which include effects on mesenchymal stem cells, angiogenesis, chondrogenesis, bone formation and resorption. Furthermore, PTH_1-34 has been shown to enhance bone repair in challenged animal models of aging, inflammatory arthritis and glucocorticoid-induced bone loss. This pre-clinical success has led to off-label clinical use and a number of case reports documenting PTH_1-34 treatment of delayed-unions and non-unions have been published. Although a recently completed phase 2 clinical trial of PTH_1-34 treatment of patients with radius fracture has failed to achieve its primary outcome, largely because of effective healing in the placebo group, several secondary outcomes are statistically significant, highlighting important issues concerning the appropriate patient population for PTH_1-34 therapy in skeletal repair. Here, we review our current knowledge of the effects of PTH_1-34 therapy for bone healing, enumerate several critical unresolved issues (e.g., appropriate dosing regimen and indications) and discuss the long-term potential of this drug as an adjuvant for endogenous tissue engineering.     

Protein Kinase PKR Mediates the Apoptosis Induction and Growth Restriction Phenotypes of C Protein-Deficient Measles Virus

The measles virus (MV) accessory proteins V and C play important roles in MV replication and pathogenesis. Infection with recombinant MV lacking either V or C causes more cell death than infection with the parental vaccine-equivalent virus (MVvac), and C-deficient virus grows poorly relative to the parental virus. Here, we show that a major effector of the C phenotype is the RNA-dependent protein kinase PKR. Using human HeLa cells stably deficient in PKR as a result of RNA interference-mediated knockdown (PKR^kd cells), we demonstrated that a reduction in PKR partially rescued the growth defect of C knockout (C^ko) virus but had no effect on the growth of either wild-type (WT) or V knockout (V^ko) virus. Increased growth of the C^ko virus in PKR^kd cells correlated with increased viral protein expression, while defective growth and decreased protein expression in PKR-sufficient cells correlated with increased phosphorylation of PKR and the α subunit of eukaryotic initiation factor 2. Furthermore, infection with WT, V^ko, or especially C^ko virus caused significantly less apoptosis in PKR^kd cells than in PKR-sufficient cells. Although apoptosis induced by C^ko virus infection in PKR-sufficient cells was blocked by a caspase antagonist, the growth of C^ko virus was not restored to the WT level by treatment with this pharmacologic inhibitor. Taken together, these results indicate that PKR plays an important antiviral role during MV infection but that the virus growth restriction by PKR is not dependent upon the induction of apoptosis. Furthermore, the results establish that a principal function of the MV C protein is to antagonize the proapoptotic and antiviral activities of PKR.     

Effect of various methods of immobilization on the stability of a microbial biosensor for surfactants based onPseudomonas rathonis T

The operating and storage stability of a receptor element of an amperometric biosensor based on thePseudomonas rathonis strain T capable of degrading surfactants was tested. Microbial cells were immobilized by incorporation in gels (agar, agarose, and calcium-alginate), polyvinyl alcohol membrane, adhesion to Chromatographic paper GF/A, or by cross-linking induced by glutaric aldehyde. Incorporation of microbial cells in agar gel provides long-standing conservation of their activity and viability during measurements of high concentrations of surfactants and allows the receptor element of the biosensor to be rapidly recovered after measurements.     

Distribution of matter in the current-carrying plasma and dense core of the discharge channel formed upon electrical wire explosion

Distribution of matter in the discharge channel formed upon a nanosecond electrical explosion of a single wire in air and vacuum was studied experimentally. Simultaneous use of optical, UV, and X-ray diagnostics made it possible to distinguish qualitatively different regions of the discharge channel, such as the current-carrying layers and the region occupied by a weakly conducting cold plasma. Several series of experiments with 25-μm-diameter 12-mm-long wires made of different materials were performed. The charging voltage and the current amplitude were varied in the ranges of U ₀ = 10–20 kV and I _max ∼ 5–10 kA, respectively. Explosion regimes with a current pause and with and without current interruption, as well as with wire preheating in air and vacuum, were studied. Shadow and schlieren images of the discharge channel were obtained using optical probing at the second harmonic of a YAG: Nd⁺³ laser (λ = 0.532 μm, τ ∼ 10 ns). In the experiments carried out in vacuum, X-ray images of the discharge channel were also obtained using an X-pinch as a point source of probing radiation and UV images were recorded using a four-frame MCP camera.     

Suppression of A-type potassium current in pyramidal neurons of the rat hippocampus, induced by pinacidil and its fluorine derivatives

With the use of a patch-clamp technique in the whole-cell configuration, we studied the effects of pinacidil and its fluorine derivatives on A-type potassium current (I _A) through the membrane of pyramidal neurons of the rat hippocampus. Hydrogen peroxide (10 mM) exerted no influence on the rate of inactivation ofI _A; therefore, this current is probably mediated by Shal Kv4.2 potassium channels. Pinacidil demonstrated the properties of a weakI _A blocker: in the 500 μM concentration it blocked about 45% of the current, while 50 μM of pinacidil fluorine derivatives were capable of blocking up to 30% ofI _A. The effects of pinacidil and its derivatives showed no dependence on the stimulating potential. A similar pattern of the effects of pinacidil fluorine derivatives, which are an order of magnitude stronger than those of pinacidil itself, allows us to suppose that the imine nitrogen of the tested compounds is significantly more involved in the molecular interaction with the site of an A-type potassium channel than the pyridine nitrogen.     

Validation of DNA methylation profiling in formalin-fixed paraffin-embedded samples using the Infinium HumanMethylation450 Microarray

A formalin-fixed paraffin-embedded (FFPE) sample usually yields highly degraded DNA, which limits the use of techniques requiring high-quality DNA, such as Infinium Methylation microarrays. To overcome this restriction, we have applied an FFPE restoration procedure consisting of DNA repair and ligation processes in a set of paired fresh-frozen (FF) and FFPE samples. We validated the FFPE results in comparison with matched FF samples, enabling us to use FFPE samples on the Infinium HumanMethylation450 Methylation array.     

"Pertussis toxin induces tachycardia and impairs the increase in blood pressure produced by alpha 2-adrenergic agonists"

Administration of purified pertussis toxin to rats induced persistent tachycardia, (observed in conscious rats but not after pithing); as little as 0.05 microgram/100 g produced a significant effect. Pertussis toxin-treatment did not affected the pressor response produced in the pithed rats by the alpha 2-adrenergic agonist methoxamine but markedly diminished the pressor effect of the alpha 2-adrenergic agonists clonidine and azepexole. A role of adenylate cyclase inhibition in the action of postsynaptic vascular alpha 2-adrenergic receptors is suggested.