Evaluation of bone healing in femurs lengthened via the gradual distraction method

BACKGROUND: Treatment of leg length inequality via lengthening of the shorter extremity is an infrequent orthopedic procedure due to the requirement of special distraction devices and possible serious complications. Essential qualitative changes in operative technique development are associated with the name of G. A. Ilizarov, who paved the way for the autoregenerate gradual distraction method in the 1950s. MATERIAL AND METHODS: In the years 1990 through 2006 a total of 57 patients underwent femur lengthening via gradual distraction using various types of external fixators at the Department of Pediatric Surgery, Orthopedics, and Traumatology, Faculty Hospital in Brno. The quality of bone healing was monitored and a number of parameters followed and statistically evaluated using regularly scheduled X-ray examinations. RESULTS: In 11 cases we had to remove the external fixator following the distraction phase, perform an osteosynthesis via a splint and fill the distraction gap via spongioplasty. The bone healing was satisfactory in the remaining 46 patients and the lengthened bone required no other fixation method. The analysis showed statistically significant deceleration in bone healing following distraction in female patients over 12 years of age, and in boys over 14 years of age. Lack of periosteal callus five weeks after surgery always signified serious problems in further healing. Severe complications were recorded in 11 cases during the distraction phase, and in 9 cases after the removal of the distraction apparatus. CONCLUSIONS: The aim of this report was to present the results of our study of distraction gap bone healing using the gradual lengthening approach.     

AIF-mediated caspase-independent necroptosis: a new chance for targeted therapeutics

Cell death has been initially divided into apoptosis, in which the cell plays an active role, and necrosis, which is considered a passive cell death program. Intense research performed in the last decades has concluded that "programmed" cell death (PCD) is a more complex physiological process than initially thought. Indeed, although in most cases the PCD process is achieved via a family of Cys proteases known as caspases, an important number of regulated PCD pathways do not involve this family of proteases. As a consequence, active forms of PCD are initially referred to as caspase-dependent and caspase-independent. More recent data has revealed that there are also active caspase-independent necrotic pathways defined as necroptosis (programmed necrosis). The existence of necroptotic forms of death was corroborated by the discovery of key executioners such as the kinase RIP1 or the mitochondrial protein apoptosis-inducing factor (AIF). AIF is a Janus protein with a redox activity in the mitochondria and a pro-apoptotic function in the nucleus. We have recently described a particular form of AIF-mediated caspase-independent necroptosis that also implicates other molecules such as PARP-1, calpains, Bax, Bcl-2, histone H2AX, and cyclophilin A. From a therapeutic point of view, the unraveling of this new form of PCD poses a question: is it possible to modulate this necroptotic pathway independently of the classical apoptotic paths? Because the answer is yes, a wider understanding of AIF-mediated necroptosis could theoretically pave the way for the development of new drugs that modulate PCD. To this end, we present here an overview of the current knowledge of AIF and AIF-mediated necroptosis. We also summarize the state of the art in some of the most interesting therapeutic strategies that could target AIF or the AIF-mediated necroptotic pathway.     

Activation of PKR causes amyloid ß-peptide accumulation via de-repression of BACE1 expression

BACE1 is a key enzyme involved in the production of amyloid ß-peptide (Aß) in Alzheimer''s disease (AD) brains. Normally, its expression is constitutively inhibited due to the presence of the 5′untranslated region (5′UTR) in the BACE1 promoter. BACE1 expression is activated by phosphorylation of the eukaryotic initiation factor (eIF)2-alpha, which reverses the inhibitory effect exerted by BACE1 5′UTR. There are four kinases associated with different types of stress that could phosphorylate eIF2-alpha. Here we focus on the double-stranded (ds) RNA-activated protein kinase (PKR). PKR is activated during viral infection, including that of herpes simplex virus type 1 (HSV1), a virus suggested to be implicated in the development of AD, acting when present in brains of carriers of the type 4 allele of the apolipoprotein E gene. HSV1 is a dsDNA virus but it has genes on both strands of the genome, and from these genes complementary RNA molecules are transcribed. These could activate BACE1 expression by the PKR pathway. Here we demonstrate in HSV1-infected neuroblastoma cells, and in peripheral nervous tissue from HSV1-infected mice, that HSV1 activates PKR. Cloning BACE1 5′UTR upstream of a luciferase (luc) gene confirmed its inhibitory effect, which can be prevented by salubrinal, an inhibitor of the eIF2-alpha phosphatase PP1c. Treatment with the dsRNA analog poly (I∶C) mimicked the stimulatory effect exerted by salubrinal over BACE1 translation in the 5′UTR-luc construct and increased Aß production in HEK-APPsw cells. Summarizing, our data suggest that PKR activated in brain by HSV1 could play an important role in the development of AD.     

Involvement of small-scale dairy farms in an industrial supply chain: when production standards meet farm diversity

In certain contexts, dairy firms are supplied by small-scale family farms. Firms provide a set of technical and economic recommendations meant to help farmers meet their requirements in terms of the quantity and quality of milk collected. This study analyzes how such recommendations may be adopted by studying six farms in Brazil. All farms are beneficiaries of the country's agrarian reforms, but they differ in terms of how they developed their activities, their resources and their milk collection objectives. First, we built a technical and economic benchmark farm based on recommendations from a dairy firm and farmer advisory institutions. Our analysis of the farms' practices and technical and economic results show that none of the farms in the sample apply all of the benchmark recommendations; however, all farms specialized in dairy production observe the main underlying principles with regard to feeding systems and breeding. The decisive factors in whether the benchmark is adopted and successfully implemented are (i) access to the supply chain when a farmer establishes his activity, (ii) a grasp of reproduction and forage production techniques and (iii) an understanding of dairy cattle feed dietary rationing principles. The technical problems observed in some cases impact the farms' dairy performance and cash position; this can lead to a process of disinvestment. This dynamic of farms facing production standards suggests that the diversity of specialized livestock farmers should be taken into account more effectively through advisory approaches that combine basic zootechnical training with assistance in planning farm activities over the short and medium term.     

Jumping without using legs: the jump of the click-beetles (Elateridae) is morphologically constrained

To return to their feet, inverted click-beetles (Elateridae) jump without using their legs. When a beetle is resting on its dorsal side, a hinge mechanism is locked to store elastic energy in the body and releases it abruptly to launch the beetle into the air. While the functional morphology of the jumping mechanism is well known, the level of control that the beetle has over this jumping technique and the mechanical constraints governing the jumps are not entirely clear. Here we show that while body rotations in air are highly variable, the jumps are morphologically constrained to a constant “takeoff” angle (79.9°±1.56°, n = 9 beetles) that directs 98% of the jumping force vertically against gravity. A physical-mathematical model of the jumping action, combined with measurements from live beetle, imply that the beetle may control the speed at takeoff but not the jumping angle. In addition, the model shows that very subtle changes in the exact point of contact with the ground can explain the vigorous rotations of the body seen while the beetle is airborne. These findings suggest that the evolution of this unique non-legged jumping mechanism resulted in a jumping technique that is capable of launching the body high into the air but it is too constrained and unstable to allow control of body orientation at landing.     

Parasitoid wasp affects metabolism of cockroach host to favor food preservation for its offspring

Unlike predators, which immediately consume their prey, parasitoid wasps incapacitate their prey to provide a food supply for their offspring. We have examined the effects of the venom of the parasitoid wasp Ampulex compressa on the metabolism of its cockroach prey. This wasp stings into the brain of the cockroach causing hypokinesia. We first established that larval development, from egg laying to pupation, lasts about 8 days. During this period, the metabolism of the stung cockroach slows down, as measured by a decrease in oxygen consumption. Similar decreases in oxygen consumption occurred after pharmacologically induced paralysis or after removing descending input from the head ganglia by severing the neck connectives. However, neither of these two groups of cockroaches survived more than six days, while 90% of stung cockroaches survived at least this long. In addition, cockroaches with severed neck connectives lost significantly more body mass, mainly due to dehydration. Hence, the sting of A. compressa not only renders the cockroach prey helplessly submissive, but also changes its metabolism to sustain more nutrients for the developing larva. This metabolic manipulation is subtler than the complete removal of descending input from the head ganglia, since it leaves some physiological processes, such as water retention, intact.     

Diversity of halophilic archaea in the crystallizers of an Adriatic solar saltern

Haloarchaeal diversity in the crystallizers of Adriatic Secovlje salterns was investigated using gene fragments encoding 16S rRNA and bacteriorhodopsin as molecular markers. Screening of 180 clones from five gene libraries constructed for each gene targeted revealed 15 different 16S rRNA and 10 different bacteriorhodopsin phylotypes, indicating higher haloarchaeal diversity than previously reported in such hypersaline environments. Furthermore, results of rarefaction analysis indicated that analysis of an increasing number of clones would have revealed additional diversity. Finally, most sequences from the crystallizers grouped within the Halorubrum branch, whereas square-shaped 'Haloquadratum' relatives, repeatedly reported to dominate crystallizer communities, were rare. Presence of such special and diverse haloarchaeal community could be attributed to the Secovlje salterns rare continuous short-cycling salt production mechanism.     

Difference in mass analysis using labeled lysines (DIMAL-K): a new, efficient proteomic quantification method applied to the analysis of astrocytic secretomes

Here we describe an original strategy for unbiased quantification of protein expression called difference in mass analysis using labeled lysine (K) (DIMAL-K). DIMAL-K is based on the differential predigestion labeling of lysine residues in complex protein mixtures. The method is relevant for proteomic analysis by two-dimensional electrophoresis and MALDI-TOF mass spectrometry. Protein labeling on lysine residues uses two closely related chemical reagents, S-methyl thioacetimidate and S-methyl thiopropionimidate. Using protein standards, we demonstrated that 1) the chemical labeling was quantitative, specific, and rapid; 2) the differentially labeled proteins co-migrated on two-dimensional gels; and 3) the identification by mass fingerprinting and the relative quantification of the proteins were possible from a single MALDI-TOF mass spectrum. The power of the method was tested by comparing and quantifying the secretion of proteins in normal and proinflammatory astrocytic secretomes (20 microg). We showed that DIMAL-K was more sensitive and accurate than densitometric image analysis and allowed the detection and quantification of novel proteins.