Nonsurgical treatment of Mason type II radial head fractures in athletes. A retrospective study.

OBJECTIVE: The best treatment for moderately displaced radial head fractures (Mason type II) still remains controversial. In cases of isolated fractures, there is no evidence that a fragment displacement of ≥ 2 mm gives poor results in conservatively treated fractures. PATIENTS AND METHODS: We retrospectively reviewed 52 patients (31M, 21F) affected by an isolated Mason type II fracture, treated with a long arm cast for two weeks between 2008 and 2013. All patients had practiced sports before being injured. They were all either bicyclists, or baseball, boxers, basketball, rugby, tennis or football players. The mean follow-up was 36 months. Elbow and forearm range of motion were measured. The Mayo Elbow Performance Score, the Broberg and Morrey rating system and the Disabilities of the Arm, Shoulder and Hand Score (DASH score) were analyzed. Follow-up radiographs were examined for evidence of consolidation, late displacement, early arthritis and non-unions. RESULTS: Flexion was slightly impaired in the injured limb when compared to the uninjured limb (137°± 6° versus 139°±5°) as were extension (-3°±6° versus 1°±4°, p < 0.05), supination (86°±6° versus 88°±3°), pronation (87°±4° versus 88°±6°) and valgus deviation (10°±4° versus 8°±3°, p < 0.05). 40 patients had no elbow complaints; 9 patients experienced occasional pain, 2 a mild instability of the elbow, and 4 a mild loss of grip strength. The DASH score was excellent in 48 patients (92.31%). In only 6 cases (11.53%) degenerative changes were greater in formerly injured elbows than in uninjured elbows. All patients returned to their previous sports activities. CONCLUSIONS: Isolated Mason type II fractures can have a good or excellent mid-term functional outcome even when treated conservatively.

Metabolic effects of TiO2 nanoparticles, a common component of sunscreens and cosmetics, on human keratinocytes.

The long-term health risks of nanoparticles remain poorly understood, which is a serious concern given their prevalence in the environment from increased industrial and domestic use. The extent to which such compounds contribute to cellular toxicity is unclear, and although it is known that induction of oxidative stress pathways is associated with this process, the proteins and the metabolic pathways involved with nanoparticle-mediated oxidative stress and toxicity are largely unknown. To investigate this problem further, the effect of TiO2 on the HaCaT human keratinocyte cell line was examined. The data show that although TiO2 does not affect cell cycle phase distribution, nor cell death, these nanoparticles have a considerable and rapid effect on mitochondrial function. Metabolic analysis was performed to identify 268 metabolites of the specific pathways involved and 85 biochemical metabolites were found to be significantly altered, many of which are known to be associated with the cellular stress response. Importantly, the uptake of nanoparticles into the cultured cells was restricted to phagosomes, TiO2 nanoparticles did not enter into the nucleus or any other cytoplasmic organelle. No other morphological changes were detected after 24-h exposure consistent with a specific role of mitochondria in this response.

miR-143 regulates hexokinase 2 expression in cancer cells.

Tumor cells activate pathways that facilitate and stimulate glycolysis even in the presence of adequate levels of oxygen in order to satisfy their continuous need of molecules, such as nucleotides, ATP and fatty acids, necessary to support their rapid proliferation. Accordingly, a variety of human tumors are characterized by elevated expression levels of the hexokinase 2 isoform (HK2). Although different molecular mechanisms, including genetic and epigenetic mechanisms, have been suggested to account for the altered expression of HK2 in tumors, the potential role of microRNAs (miRNAs) in the regulation of HK2 expression has not been evaluated. Here, we report that miR-143 inhibits HK2 expression via a conserved miR-143 recognition motif located in the 3'-untranslated region (3'UTR) of HK2 mRNA. We demonstrate that miR143 inhibits HK2 expression both in primary keratinocytes and in head and neck squamous cell carcinoma (HNSCC)-derived cell lines. Importantly, we found that miR-143 inversely correlates with HK2 expression in HNSCC-derived cell lines and in primary tumors. We also report that the miRNA-dependent regulation of hexokinase expression is not limited to HK2 as miR-138 targets HK1 via a specific recognition motif located in its 3'UTR. All these data unveil a new miRNA-dependent mechanism of regulation of hexokinase expression potentially important in the regulation of glucose metabolism of cancer cells.

Endocannabinoid signaling in Alzheimer’s disease: current knowledge and future directions.

The importance of the endocannabinoid system (ECS) in the modulation functions of the central nervous system has been extensively investigated during the last few years. In particular, accumulated evidence has implicated ECS in the pathophysiology of Alzheimer’s disease (AD), that is a progressive, degenerative, and irreversible disorder characterized by the accumulation in the brain of beta-amyloid fragments forming insoluble plaques, and of intracellular neurofibrillary tangles (NTFs) associated with synaptic and neuronal loss. In all the processes involved in the formation of both plaques and NFTs, the key-role played by the ECS has been documented. Here, we review current knowledge and future directions of ECS modulation both in animal models of AD and in human tissues, underlying the role of endocannabinoid signaling in the development of AD hallmarks. Overall, the available data suggest that next generation therapeutics might target distinct ECS elements, for instance CB2 receptor or fatty acid amide hydrolase, as a promising approach to halt or at least to slow down disease progression.

Cannabinoid receptor signalling in neurodegenerative diseases: a potential role for membrane fluidity disturbance.

Type-1 cannabinoid receptor (CB(1)) is the most abundant G-protein-coupled receptor (GPCR) in the brain. CB(1) and its endogenous agonists, the so-called 'endocannabinoids (eCBs)', belong to an ancient neurosignalling system that plays important functions in neurodegenerative and neuroinflammatory disorders like Alzheimer's disease, Parkinson's disease, amyotrophic lateral sclerosis and multiple sclerosis. For this reason, research on the therapeutic potential of drugs modulating the endogenous tone of eCBs is very intense. Several GPCRs reside within subdomains of the plasma membranes that contain high concentrations of cholesterol: the lipid rafts. Here, the hypothesis that changes in membrane fluidity alter function of the endocannabinoid system, as well as progression of particular neurodegenerative diseases, is described. To this end, the impact of membrane cholesterol on membrane properties and hence on neurodegenerative diseases, as well as on CB(1) signalling in vitro and on CB(1) -dependent neurotransmission within the striatum, is discussed. Overall, present evidence points to the membrane environment as a critical regulator of signal transduction triggered by CB(1) , and calls for further studies aimed at better clarifying the contribution of membrane lipids to eCBs signalling. The results of these investigations might be exploited also for the development of novel therapeutics able to combat disorders associated with abnormal activity of CB(1).

Biomechanical evaluation of different anterior cruciate ligament fixation techniques for hamstring graft.

BACKGROUND: A number of anterior cruciate ligament (ACL) fixation techniques are currently in use. Slippage or failure of the graft by excessive loading or aggressive rehabilitation may result in an unstable knee. Load and slippage of the ACL graft varies according to the fixation technique used. METHODS: Graft slippage, load to failure, and stiffness were evaluated using an animal model. Six soft tissue ACL fixation techniques and bone cement as a fixation device were tested: group A, Endo Button CL-Bio RCI; group B, Swing Bridge-Evolgate; group C, Rigidfix-Intrafix; group D, Bone Mulch-Washer Lock; group E, Transfix-Retroscrew; group F, Transfix-Deltascrew; group G, Kryptonite bone cement. Maximum failure load, stiffness, and slippage at the 1st and 1000th cycles and mode of failure were evaluated. RESULTS: The maximum failure load was significantly higher in group B (1030 N) and significantly lower in group E (483 N) than in the others. The stiffness of group B (270 N/mm) was significantly higher than the others. As for the mode of failure, group C showed failure in the femoral side in all tests (four device ruptures and two tendon ruptures on the femoral side). All failures of the other groups occurred on the tibial side except one test in group A. All failures in group G were due to slippage of the tendons. CONCLUSION: Load to failure and stiffness was significantly different between the ACL fixation techniques. All but one of the fixation techniques showed sufficient properties for adequate postoperative rehabilitation. Bone cement used as a fixation device in soft tissue grafts did not seem to provide adequate initial fixation suitable for early rehabilitation after ACL reconstruction.

Evidence for the intracellular accumulation of anandamide in adiposomes.

Anandamide is a lipid messenger that carries out a wide variety of biological functions. It has been suggested that anandamide accumulation involves binding to a saturable cellular component. To identify the structure(s) involved in this process, we analyzed the intracellular distribution of both biotinylated and radiolabeled anandamide, providing direct evidence that lipid droplets, also known as adiposomes, constitute a dynamic reservoir for the sequestration of anandamide. In addition, confocal microscopy and biochemical studies revealed that the anandamide-hydrolase is also spatially associated with lipid droplets, and that cells with a larger adiposome compartment have an enhanced catabolism of anandamide. Overall, these findings suggest that adiposomes may have a critical role in accumulating anandamide, possibly by connecting plasma membrane to internal organelles along the metabolic route of this endocannabinoid.

Endocannabinoids in adipocytes during differentiation and their role in glucose uptake.

The molecular basis for the control of energy balance by the endocannabinoid anandamide (AEA) is still unclear. Here, we show that murine 3T3-L1 fibroblasts have the machinery to bind, synthesize and degrade AEA, and that their differentiation into adipocytes increases by approximately twofold the binding efficiency of cannabinoid receptors (CBR), and by approximately twofold and approximately threefold, respectively, the catalytic efficiency of the AEA transporter and AEA hydrolase. In contrast, the activity of the AEA synthetase and the binding efficiency of vanilloid receptor were not affected by the differentiation process. In addition, we demonstrate that AEA increases by approximately twofold insulin-stimulated glucose uptake in differentiated adipocytes, according to a CB1R-dependent mechanism that involves nitric oxide synthase, but not lipoxygenase or cyclooxygenase. We also show that AEA binding to peroxisome proliferator-activated receptor-gamma, known to induce differentiation of 3T3-L1 fibroblasts into adipocytes, is not involved in the stimulation of glucose uptake.

Biochemical changes in endocannabinoid system are expressed in platelets of female but not male migraineurs.

The endogenous cannabinoid anandamide (AEA) plays important roles in modulating pain. Head pain is an almost universal human experience, yet primary headache disorders, such as migraine without aura (MoA) or episodic tension-type headache (ETTH), can represent a serious threat to well-being when frequent and disabling. We assessed the discriminating role of endocannabinoids among patients with ETTH or MoA, and control subjects. We measured the activity of AEA hydrolase and AEA transporter, and the level of cannabinoid receptors in peripheral platelets from MoA, ETTH and healthy controls. Sixty-nine headache patients and 36 controls were selected. Diagnosis of headache type was made according to the International Headache Society criteria. We observed significant sex differences concerning AEA membrane transporter and fatty acid amide hydrolase activity in all groups. An increase in the activity of AEA hydrolase and AEA transporter was found in female but not male migraineurs. Cannabinoid receptors were the same in all groups. Here we show that the endocannabinoid system in human platelets is altered in female but not male migraneurs. Our results suggest that in migraineur women an increased AEA degradation by platelets, and hence a reduced concentration of AEA in blood, might reduce the pain threshold and possibly explain the prevalence of migraine in women. The involvement of the endocannabinoid system in migraine is new and broadens our knowledge of this widespread and multifactorial disease.

Up-regulation of the endocannabinoid system in the uterus of leptin knockout (ob/ob) mice and implications for fertility.

The levels of the endocannabinoids anandamide and 2-arachidonoylglycerol (2-AG) are under the negative control of leptin in the rodent hypothalamus. As leptin and endocannabinoids play opposite roles in the control of reproduction, we have investigated whether the impaired fertility typical of leptin-defective ob/ob mice is due, in part, to enhanced uterine endocannabinoid levels. We found that levels of both anandamide and 2-AG in the uterus of ob/ob mice are significantly elevated with respect to wild-type littermates, due to reduced hydrolase activity in the case of anandamide, and to reduced monoacylglycerol lipase and enhanced diacylglycerol lipase activity in the case of 2-AG. Furthermore, the process mediating endocannabinoid cellular uptake was also impaired in ob/ob mice, whereas the levels of cannabinoid and anandamide receptors were not modified. Although ineffective in wild-type mice, treatment of ob/ob mice with leptin re-established endocannabinoid levels and enzyme activities back to the values observed in wild-type littermates. Finally, treatment of ob/ob females with the CB1 receptor antagonist SR141716A did not improve their fertility, and inhibition of endocannabinoid inactivation with the endocannabinoid uptake inhibitor OMDM-1 in wild-type females did not result in impaired fertility.

Transglutaminase 5 is acetylated at the N-terminal end.

Transglutaminases (TGases) are calcium-dependent enzymes that catalyse cross-linking between proteins by acyl transfer reaction; they are involved in many biological processes including coagulation, differentiation, and tissue repair. Transglutaminase 5 was originally cloned from keratinocytes, and a partial biochemical characterisation showed its involvement in skin differentiation, in parallel to TGase 1 and TGase 3. Here, we demonstrate, by electrospray tandem mass spectrometry that TGase 5 is acetylated at the N-terminal end. Moreover, in situ measurement of TGase activity shows that endogenous TGase 5 is active upon treatment with phorbol acetate, and the enzyme co-localises with vimentin intermediate filaments.

Fluctuations of fatty acid amide hydrolase and anandamide levels during the human ovulatory cycle.

Implantation is possible within a defined period of the menstrual cycle, referred to as the 'implantation window'. It is during this critical period that proper dialog can be established between the blastocyst and a receptive endometrium. If for any reason this dialog is not established or is altered, the embryo is aborted. The factors responsible for the interaction between the embryo and the mother at the moment of implantation remain poorly understood. Recent studies indicate that endocannabinoids may contribute to the development of an adequate milieu at the implantation site. Here we show that the levels of anandamide and of its degrading enzyme, the fatty acid amide hydrolase, in peripheral lymphocytes undergo specific variations during the various phases of the human ovulatory cycle. In particular, we found the highest levels of fatty acid amide hydrolase activity and protein content, paralleled by the lowest anandamide concentrations, in the period that temporally coincides with the putative window of implantation in humans. On the other hand, the anandamide-synthesizing phospholipase D, the anandamide membrane transporter and the anandamide-binding cannabinoid receptors of lymphocytes did not change during the menstrual cycle. This study indicates that high fatty acid amide hydrolase activity and low anandamide levels may be among the factors that co-operate in the success of implantation. This would add to our understanding of the pathophysiological and therapeutic implications of the endocannabinoid system in human fertility.

Mouse blastocysts release a lipid which activates anandamide hydrolase in intact uterus.

Anandamide (N-arachidonoylethanolamine, AEA) is a major endocannabinoid, known to impair mouse pregnancy and embryo development and to induce apoptosis in blastocysts. Here we show that mouse blastocysts rapidly (within 30 min of culture) release a soluble compound, that increases by approximately 2.5-fold the activity of AEA hydrolase (fatty acid amide hydrolase, FAAH) present in the mouse uterus, without affecting FAAH gene expression at the translational level. This "FAAH activator" was produced by both trophoblast and inner cell mass cells, and its initial biochemical characterization showed that it was fully neutralized by adding lipase to the blastocyst-conditioned medium (BCM), and was potentiated by adding trypsin to BCM. Other proteases, phospholipases A(2), C or D, DNAse I or RNAse A were ineffective. BCM did not affect the AEA-synthesizing phospholipase D, the AEA-binding cannabinoid receptors, or the selective AEA membrane transporter in mouse uterus. The FAAH activator was absent in uterine fluid from pregnant mice and could not be identified with any factor known to be released by blastocysts. In fact, platelet-activating factor inhibited non-competitively FAAH in mouse uterus extracts, but not in intact uterine horns, whereas leukotriene B(4) or prostaglandins E(2) and F(2)alpha had no effect. Overall, it can be suggested that blastocysts may protect themselves against the noxious effects of uterine endocannabinoids by locally releasing a lipid able to cross the cell membranes and to activate FAAH. The precise molecular identity of this activator, the first ever reported for FAAH, remains to be elucidated.

The endocannabinoid system, anandamide and the regulation of mammalian cell apoptosis.

Endocannabinoids are a new class of lipid mediators, which include amides, esters and ethers of long-chain polyunsaturated fatty acids. Anandamide (N-arachidonoylethanolamine; AEA) and 2-arachidonoylglycerol (2-AG) are the main endogenous agonists of cannabinoid receptors able to mimic several pharmacological effects of Delta-9-tetrahydrocannabinol, the active principle of Cannabis sativa preparations like hashish and marijuana. The pathways leading to the synthesis and release of AEA and 2-AG from neuronal and non-neuronal cells are still rather uncertain. Instead, it is known that the activity of AEA is limited by cellular uptake through a specific membrane transporter, followed by intracellular degradation by a fatty acid amide hydrolase. Together with AEA and congeners these proteins form the 'endocannabinoid system'. Here, the involvement of AEA in apoptosis and the underlying signal transduction pathways will be reviewed, along with the metabolic routes and the molecular targets of this endocannabinoid. Also, recent findings on the apoptotic potential of AEA for neuronal cell differentiation and brain development will be discussed.

Endocannabinoids, hormone-cytokine networks and human fertility.

Anandamide (N -arachidonoylethanolamine, AEA) is a major endocannabinoid, shown to impair mouse pregnancy and embryo development and to induce apoptosis in blastocysts. Here, we review the roles of AEA, of the AEA-binding cannabinoid (CB) receptors, of the selective AEA membrane transporter (AMT), and of the AEA-hydrolyzing enzyme fatty acid amide hydrolase (FAAH), in human gestation. In particular, we discuss the interplay between the endocannabinoid system and the hormone-cytokine array involved in the control of human pregnancy, showing that the endocannabinoids take part in the immunological adaptation occurring during early pregnancy. In this line, we discuss the critical role of FAAH in human peripheral lymphocytes, showing that the expression of this enzyme is regulated by progesterone, Th1 and Th2 cytokines, which also regulate fertility. Moreover, we show that AEA and the other endocannabinoid, 2-arachidonoylglycerol, inhibit the release of the fertility-promoting cytokine leukemia inhibitory factor from human lymphocytes. Taken together, low FAAH and consistently high blood levels of AEA, but not CB receptors or AMT, can be early (<8 weeks of gestation) markers of spontaneous abortion, potentially useful as diagnostic tools for large-scale, routine monitoring of gestation in humans.

d-penicillamine reduces serum oxidative stress in Alzheimer's disease patients.

BACKGROUND: Several lines of evidence address the emerging role for copper in Alzheimer's disease (AD) for sustaining oxidative mechanisms. Studies indicate that peripheral markers of oxidative stress in AD patients could be informative about the pathophysiology of this brain condition. Here, we present a pilot study examining the efficacy of the copper-chelating agent d-penicillamine in reducing oxidative stress in AD patients. DESIGN: Serum levels of copper sampled in AD patients and healthy controls indicate a copper homeostasis imbalance in AD. On this basis, 34 AD patients were enrolled in a 6-month, double-blind, placebo-controlled trial with the copper d-penicillamine-chelating agent. Nine patients for each group completed the trial. Oxidative stress, trace metals and clinical parameters were evaluated. RESULTS: At the start of the study (t0) total peroxides and copper serum content of AD patients were higher (P < 0.0001, P < 0.0001, respectively) and antioxidants were lower (P < 0.05) than in healthy controls. Copper and peroxides were correlated in the AD population (Pearson's r = 0.61, P < 0.001). After treatment with d-penicillamine, the extent of oxidative stress (P < 0.05) was decreased, but no difference was observed in the rate of cognitive decline. CONCLUSION: Data from this pilot study suggest that copper could play a role in the production of peroxides in AD, and that d-penicillamine has an effect in reducing oxidative damage, however, results are still inconclusive in terms of drug efficacy on the clinical progression of AD. Studies with larger cohorts are needed to elucidate the real effectiveness of d-penicillamine treatment in AD.