Vigna unguiculata L. Walp. Leaves as a Source of Phytochemicals of Dietary Interest: Optimization of Ultrasound-Assisted Extraction and Assessment of Traditional Consumer Habits.

Vigna unguiculata L. Walp. is an African crop spread worldwide mainly for pulses production. Despite being a neglected and under-utilized food, cowpea leaves are a rich source of phytochemicals and micronutrients. The aim of the work is to characterize the phytochemical composition of cowpea leaves by an optimized ultrasound-assisted extraction (USAE) and to compare raw and boiled leaves. A three-level factorial design (Box-Behnken) was employed for the optimization of the USAE considering three different parameters (% ethanol, drug-to-solvent ratio, and number of cycles). The optimized extracts were characterized by LC/MS/MS. Finally, leaves were boiled at 100 °C for 30 min to simulate traditional cooking procedures and compared to raw leaves. The best extraction condition was EtOH/H2 O 1 : 2 v/v, drug to solvent ratio 1 : 47 w/v, and 3 extraction cycles. The phytochemicals identified mainly belong to the family of phenolic acids, flavonoids, terpenoids, and alkaloids. Boiled leaves revealed a significant loss of most phytochemicals and a net decrease of their antioxidant activity compared to the raw ones. The results highlight the potential nutraceutical value of cowpea leaves whilst the impoverishment triggered by traditional consumer habits pushes the need to evaluate alternative cooking procedures helpful in the maintenance of their phytochemical properties.

Multidetection scheme for transient-grating-based spectroscopy.

Time-resolved optical spectroscopy represents an effective non-invasive approach to investigate the interplay of different degrees of freedom, which plays a key role in the development of novel functional materials. Here, we present magneto-acoustic data on Ni thin films on SiO2 as obtained by a versatile pump-probe setup that combines transient grating spectroscopy with time-resolved magnetic polarimetry. The possibility to easily switch from a pulsed to continuous wave probe allows probing of acoustic and magnetization dynamics on a broad time scale, in both transmission and reflection geometry.

Prenylcysteine oxidase 1, an emerging player in atherosclerosis.

The research into the pathophysiology of atherosclerosis has considerably increased our understanding of the disease complexity, but still many questions remain unanswered, both mechanistically and pharmacologically. Here, we provided evidence that the pro-oxidant enzyme Prenylcysteine Oxidase 1 (PCYOX1), in the human atherosclerotic lesions, is both synthesized locally and transported within the subintimal space by proatherogenic lipoproteins accumulating in the arterial wall during atherogenesis. Further, Pcyox1 deficiency in Apoe-/- mice retards atheroprogression, is associated with decreased features of lesion vulnerability and lower levels of lipid peroxidation, reduces plasma lipid levels and inflammation. PCYOX1 silencing in vitro affects the cellular proteome by influencing multiple functions related to inflammation, oxidative stress, and platelet adhesion. Collectively, these findings identify the pro-oxidant enzyme PCYOX1 as an emerging player in atherogenesis and, therefore, understanding the biology and mechanisms of all functions of this unique enzyme is likely to provide additional therapeutic opportunities in addressing atherosclerosis.

Direct anterior approach and dual mobility cup: the "head-first" reduction technique: a technical note and early case series.

PURPOSE: Less invasive direct anterior approach (DAA) and dual mobility cup (DMC) are increasingly adopted in practice over the last decade. Their use aims to reduce, as much as possible, soft tissue dissection and dislocation rate. This study aims to present a novel surgical technique to reduce a DMC prosthesis during a DAA easily. METHODS: A mildly modified version of the direct anterior approach is proposed. When leg lengths, stability, impingement, and tension have been checked, the trial stem is disassembled in situ, dislocated, and removed, leaving the space to exchange the trial double mobility head with the definitive one. When the definitive stem is inserted, the surgeon guides and helps the assistant to match the trunnion in the double mobility head. As soon as the components are matched, the traction is released, and the unit is impacted by an alternation of axial traction and release. RESULTS: Of 164 patients who underwent primary total hip arthroplasty (December 2016-May 2017) by a single surgeon, a double mobility cup through DAA and the "head-first" technique was performed in 26 patients (15.8%). The mean operative time was 130 min (85-220 min; SD 34.28). No significant complications occurred during the mean follow-up of 23.6 months. CONCLUSIONS: Specific difficulties can be anticipated when pairing dual mobility cup and direct anterior approach. The "head-first" technique is a useful technique in reducing the possible difficulties related to the reduction of double mobility cup through a less invasive direct anterior approach.

Soft-tissue defects of the Achilles tendon region: Management and reconstructive ladder. Review of the literature.

INTRODUCTION: Defects of the Achilles tendon region represent a challenge for reconstructive surgeons. Several options are available but there is still no reconstructive ladder for this specific and tricky area. An up-to-date reconstructive ladder according to local and general conditions is proposed based on our multicentre experience and an extensive review of the English literature on PubMed. MATERIALS AND METHODS: An extensive review of the English literature was performed on PubMed using the following key-words: "Achilles region", "heel", "soft-tissue reconstruction", "flaps", "grafts" and "dermal substitutes". RESULTS: A total of 69 complete papers were selected, covering the last thirty years' literature. Although most of the studies were based on limited case-series, local and general conditions were always reported. A comprehensive reconstructive ladder of all the available reconstructive techniques for the Achilles region has been created based on our personal multicentre experience and the results of the literature review. CONCLUSIONS: The reconstructive ladder is a concept that is still a mainstay in plastic surgery and guides decisions in the repair strategy for soft tissue defects. The optimal solution, according to the experience of the surgeon and the wishes of the patient, is the one that implies less sacrifice of the donor site. Perforator flaps should be the first-line option for small-to-moderate defects; the distally-based sural flap is the most reported for moderate-to-large defects of the Achilles region, and free flaps should be reserved mainly for complex and wide reconstructions.

Data for proteomic analysis of Human monocyte-derived macrophages.

This data article is referred to the research article entitled Human monocyte-derived macrophages are heterogeneous: proteomic profile of different phenotypes by Eligini et al. Eligini S., Brioschi M., Fiorelli S., Tremoli E., Banfi C., Colli S. Human monocyte-derived macrophages are heterogeneous: proteomic profile of different phenotypes. J. Proteomics 124, 2015, 112-123. Macrophages obtained in vitro from blood monocytes are largely used as surrogate model of tissue macrophages that are heterogeneous and not easy to obtain and handle. Under spontaneous differentiation in vitro, monocyte-derived macrophages (MDMs) display two dominant subsets (round and spindle) that show different transcriptional, antigenic, and functional profiles mimicking, at least in part, the heterogeneity of tissue macrophages. This article reports the nano-LC-MS(E) analysis of the proteome of round and spindle MDMs allowing a deeper comprehension of macrophage heterogeneity.

Human monocyte-derived macrophages are heterogenous: Proteomic profile of different phenotypes.

Tissue macrophages play a key role in many aspects of human physiology and pathology. These cells are heterogeneous both in term of morphology and function. As an example, heterogeneity has been reported within the atherosclerotic lesions where distinct populations exert opposite functions driving plaque progression or stability. Tissue macrophages are not easily obtained and differentiated blood-derived monocytes are largely used as surrogate model. We previously reported that human macrophages spontaneously differentiated from adherent monocytes show two dominant subsets, distinct for morphology (spindle and round) and functions. The aim of this study was to evaluate the intracellular proteome of these two macrophage subsets by means of a microproteomic workflow properly set up to simultaneously identify and quantify proteins from a minimal number of morphotypically heterogeneous cells in culture. We report two distinct proteomic profiles that distinguish round from spindle macrophages. In particular, differential abundances were observed for proteins involved in membrane traffic regulation, lipid handling, efferocytosis, and protection against stress conditions. Results reinforce and extend previous data on the functional and antigenic profile of these macrophage phenotypes strengthening the suitability of our model to focus on macrophage heterogeneity. BIOLOGICAL SIGNIFICANCE: Tissue macrophages patrol homeostatic functions, immune surveillance, and resolution of inflammation. The spectrum of macrophage activation states is, therefore, wide and gives ground for the heterogeneity of these cells, documented in health and disease. This study provides knowledge of the distinct proteome that characterises the two dominant morphotypes (round and spindle) of human macrophages that, in our culture condition, are generated by spontaneous differentiation from blood-derived monocytes. Results extend previous data about the different antigenic, transcriptional, and functional profiles of these morphotypes and further strengthen the suitability of this in vitro model to study macrophage heterogeneity and to address the effects of environmental challenges and drugs.

Posterior tibial artery perforator flaps for coverage of Achilles region defects.

BACKGROUND: Defects of the Achilles tendon region still represent a tricky issue in lower limb surgery. Among the several reconstructive possibilities, local propeller perforator flaps have gained popularity in the last decade. MATERIALS AND METHODS: We report our experience with eight patients affected by small-to-moderate soft-tissue defects of the Achilles tendon region, who underwent surgical reconstruction with local flaps based on posterior tibial perforator branches. RESULTS: All patients healed successfully in terms of aesthetic and functional aspect. In only one case a transient venous congestion was observed and this resolved spontaneously. CONCLUSIONS: Although the surgical technique requires much care and skill, including an extremely gentle dissection of perforator vessels, local propeller flaps should be considered the first-line choice for reconstruction in small-to-medium size soft-tissue defects in the Achilles region.

Thermographic characterization of masticatory muscle regions in volunteers with and without myogenous temporomandibular disorder: preliminary results.

OBJECTIVES: This study aims to conduct a non-invasive measurement of the cutaneous temperature of selected masticatory muscle regions of volunteers with and without myogenous temporomandibular disorder (TMD), using infrared thermography. METHODS: 23 females (10 myogenous TMD volunteers and 13 controls) were recruited and studied. The temperature at the surface of the facial area over the anterior temporalis and masseter muscles was assessed by medical thermography, using regional lateral views and clinical examination. RESULTS: The temperature levels measured at the masseter and anterior temporalis muscle regions in myogenous TMD volunteers (32.85 ± 0.85 and 34.37 ± 0.64 ºC, respectively) were significantly lower (p < 0.05) than those measured in controls (33.49 ± 0.92 and 34.78 ± 0.44 ºC, respectively). Medical infrared imaging indicated a mean difference of 1.4 ºC between the masseter and anterior temporalis regions. Analysis of the comparison between the absolute and normalized mean temperatures was performed using the pairwise comparison of receiver operating characteristic curves, and no statistically significant difference was observed (p > 0.05). The sensitivity and specificity of the thermographic assessment for the masseter region was of 70% and 73%, respectively and for the anterior temporalis region was of 80% and 62%, respectively. CONCLUSIONS: This method of evaluating masticatory muscle regions of this preliminary study seems to indicate that it can be used as an aid in complimentary diagnosing of TMDs.

Thermographic and clinical correlation of myofascial trigger points in the masticatory muscles.

OBJECTIVES: The aim of the study was to identify and correlate myofascial trigger points (MTPs) in the masticatory muscles, using thermography and algometry. METHODS: 26 female volunteers were recruited. The surface facial area over the masseter and anterior temporalis muscles was divided into 15 subareas on each side (n=780). This investigation consisted of three steps. The first step involved thermographic facial examination, using lateral views. The second step involved the pressure pain threshold (PPT), marking the MTP pattern areas for referred pain (n=131) and local pain (n=282) with a coloured pencil, and a photograph of the lateral face with the head in the same position as the infrared imaging. The last step was the fusion of these two images, using dedicated software (Reporter® 8.5-SP3 Professional Edition and QuickReport® 1.2, FLIR Systems, Wilsonville, OR); and the calculation of the temperature of each point. RESULTS: PPT levels measured at the points of referred pain in MTPs (1.28±0.45 kgf) were significantly lower than the points of local pain in MTPs (1.73±0.59 kgf; p<0.05). Infrared imaging indicated differences between referred and local pain in MTPs of 0.5 °C (p<0.05). Analysis of the correlation between the PPT and infrared imaging was done using the Spearman non-parametric method, in which the correlations were positive and moderate (0.4≤r<0.7). The sensitivity and specificity in MTPs were 62.5% and 71.3%, respectively, for referred pain, and 43.6% and 60.6%, respectively, for local pain. CONCLUSION: Infrared imaging measurements can provide a useful, non-invasive and non-ionizing examination for diagnosis of MTPs in masticatory muscles.

Statins prevent tissue factor induction by protease-activated receptors 1 and 2 in human umbilical vein endothelial cells in vitro.

BACKGROUND: Protease-activated receptors (PARs) are G-protein-coupled receptors that function in hemostasis and thrombosis, as well as in the inflammatory and proliferative responses triggered by tissue injury. We have previously shown that PAR1 or PAR2 occupancy by specific PAR-agonist peptides (PAR-APs) induces tissue factor (TF) expression in human umbilical vein endothelial cells (HUVECs), where TF regulation by PAR1 (but not by PAR2) requires intact endothelial caveolin-enriched membrane microdomains in which PAR1 and caveolin-1 associate. OBJECTIVES: The aim of this study was to determine the effects of cholesterol-lowering agents (statins) and cholesterol-loading lipoprotein on PAR1-AP-mediated and PAR2-AP-mediated TF induction in HUVECs. RESULTS: Statins completely prevented TF induction by PAR-APs in an isoprenoid-independent manner, induced the delocalization of PAR1 from caveolin-enriched membrane microdomains without affecting PAR1 mRNA, and decreased PAR2 mRNA and protein levels. Statins also prevented PAR-AP-mediated extracellular signal-related kinase 1/2 activation, which is crucial for TF induction. The redistribution of PAR1 is accompanied by the relocation of the membrane microdomain-associated G-protein α, caveolin-1, and Src, which we previously showed to play a key role in signal transduction and TF induction. Conversely, cholesterol loading potently amplified PAR1-AP-induced TF, probably as a result of the increased abundance of PAR1 and the Src and G-protein α signaling molecules in the caveolin-1-enriched fraction, without affecting PAR1 mRNA. CONCLUSIONS: As PARs have important functions in hemostasis, cancer, thrombosis, and inflammatory processes, our findings that statins prevent TF induction by PAR-APs altering the membrane localization of PAR1 and the expression of PAR2 suggest that they may provide health benefits other than reducing atherosclerosis.

Surfactant protein B and RAGE increases in the plasma during cardiopulmonary bypass: a pilot study.

Surfactant derived protein B (SPB) and plasma receptor for advanced glycation end products (RAGE) have been proposed as markers of lung injury. The former is produced specifically by pneumocytes while RAGE production is present in several body tissues. Cardiopulmonary bypass (CPB) generates a transient lung injury. We measured SPB and RAGE in plasma before surgery and after CPB, as well as 24 h and 48 h later. We analysed plasma samples from 20 subjects scheduled for elective coronary artery bypass grafting. We performed a quantitative analysis of plasma levels of RAGE and SPB mature form (8 kDa) by ELISA and a semi-quantitative analysis of SPB immature form (~ 40 kDa) by Western blotting. Surgery procedures were uneventful. After CPB RAGE median (75th-25th interquartile difference) increased from 633 (539) pg·mL⁻¹ to 1,362 (557) pg·mL⁻¹ (p < 0.01), while mature SPB increased from 5,587 (3,089) ng·mL⁻¹ to 20,307 (19,873) ng·mL⁻¹ (p < 0.01). RAGE and mature SPB returned to normal values within 48 h. This behaviour was confirmed when RAGE and SPB were normalised for protein content. Parallel changes were observed for immature SPB. Plasma RAGE and SPBs are sensitive and rapid markers of lung distress.

Cerebral amyloid angiopathy-related inflammation: an emerging disease.

Three elderly patients with, respectively: mild cognitive impairment, severe and progressive neurologic involvement, and focal neurologic deficit, were observed. MRI showed multiple areas of white matter edema, at times partially involving the cortex, in the first two patients, and a single area in the third. Treatment with steroids determined the disappearance of the lesions and clinical amelioration. The key to the diagnosis of cerebral amyloid angiopathy-related inflammation (CAA-ri) was the demonstration, with appropriate MRI sequences, of microbleeds consistent with cerebral amyloid angiopathy (CAA). This diagnosis was supported by genetic analysis of APOE with demonstration of ε4/ε4 genotype, found in about 80% of CAA patients who develop inflammatory changes. In the appropriate clinical setting, MRI demonstration of microbleeds supported by results of genetic analysis of APOE may strongly support the diagnosis of CAA-ri thus avoiding cerebral biopsy.

Proteomic profile of differentially expressed plasma proteins from dystrophic mice and following suberoylanilide hydroxamic acid treatment.

PURPOSE: Histone Deacetylase Inhibitors (DI) ameliorates dystrophic muscle regeneration restoring muscular strength in the mdx mouse model of Duchenne muscular dystrophy (DMD). The further development of these compounds as drugs for DMD treatment is currently hampered by the lack of knowledge about DIs effect in large dystrophic animal models and that of suitable biomarkers to monitor their efficacy. EXPERIMENTAL DESIGN: In this study we applied proteomic analysis to identify differentially expressed proteins present in plasma samples from mdx mice treated with the Suberoylanilide hydroxamic acid (SAHA) and relative normal controls (WT). RESULTS: Several differentially expressed proteins were identified between untreated wild type and mdx mice. Among these, fibrinogen, epidermal growth factor 2 receptor, major urinary protein and glutathione peroxidase 3 (GPX3) were constitutively up-regulated in mdx, while complement C3, complement C6, gelsolin, leukaemia inhibitory factor receptor (LIFr), and alpha 2 macroglobulin were down-regulated compared to WT mice. SAHA determined the normalization of LIFr and GPX3 protein level while apoliprotein E was de novo up-regulated in comparison to vehicle-treated mdx mice. CONCLUSIONS AND CLINICAL RELEVANCE: Collectively, these data unravel potential serological disease biomarkers of mdx that could be useful to monitor muscular dystrophy response to DI treatment.

Proteomic analysis of atherosclerotic plaque.

Proteins play a fundamental role in the formation and progression of plaque, but proteomic analysis of plaque as a whole is difficult, due to its heterogeneous cellular composition and an abundance of plasma proteins. Several approaches to this problem are reported in the literature; they include proteomic analysis of vascular tissues, analysis of proteins released by normal and pathological arterial walls, proteomic analysis of vascular cells and proteomic analysis of blood. In a previous study, we proposed a new strategy for studying of proteome of plaque, which permits to select the proteins exclusive to plaque by the constructing of a reference synthetic gel. In the present work, we matched the spots of the reference synthetic gel with the spots of a pool of carotid plaque, in order to select only spots exclusive to plaque from the 2-dimensional electrophoresis of the pool of plaque. We selected some spots between those exclusive and identified them by mass spectrometry. Some proteins identified are involved in transport, others take part in elimination of toxic radicals, others are metabolic enzymes or structural proteins. This study represents an example of application of the new approach which we have proposed: the reference gel of proteome of plaque permits to select, on every sample of interest, only the spots exclusive to plaque; once selected, spots can be identified by mass spectrometry and, being typical of plaque composition, could represent novel markers of lesions and vascular risk.

Mitochondrial reactive oxygen species: a common pathway for PAR1- and PAR2-mediated tissue factor induction in human endothelial cells.

BACKGROUND: Protease-activated receptors (PARs) comprise a family of G-protein-coupled receptors with a unique proteolytic activation mechanism. PARs regulate a broad range of cellular functions and are involved in the pathogenesis of inflammatory disorders. Moreover, PAR1 and PAR2 activation in the endothelium shifts it toward a prothrombotic condition. OBJECTIVES: To assess the relevance of intracellular reactive oxygen species (ROS) in the signaling events underlying tissue factor (TF) expression elicited by PAR1 and PAR2 occupancy in endothelial cells, and to investigate their source. METHODS: Human umbilical vein endothelial cells (HUVEC) were exposed to specific PAR1 and PAR2 agonist peptides. TF expression was determined by real-time reverse transcription polymerase chain reaction analysis and measurement of procoagulant activity. ROS generation was determined by a fluorometric assay after cell loading with 2'-7'-dichlorofluorescein diacetate. RESULTS: ROS generated by the mitochondrial chain, mostly from complex III, provide a pathway through which PAR1 and PAR2 occupancy induces TF. Other sources of ROS do not participate in TF induction. Activation of both ERK1/2 and p38 MAPK is critical for mitochondrial ROS generation. In addition to these pathways shared by the two PARs, mechanisms downstream from PAR1 and PAR2 activation, different for the two receptors, also induced TF. A module that sensitively regulates PAR1 signaling and ultimately involves NF-kappaB activation has been identified. CONCLUSIONS: Our data identify ROS originating in mitochondria as key mediators of the signaling pathways triggered by PAR1 and PAR2 engagement in endothelial cells and show that downstream from receptor activation occur cascades that are mechanistically coupled to procoagulant activity.

Tissue factor induction by protease-activated receptor 1 requires intact caveolin-enriched membrane microdomains in human endothelial cells.

BACKGROUND: Protease-activated receptors (PARs) comprise a family of G-protein-coupled receptors with a unique mechanism of proteolytic activation. PARs regulate a broad range of cellular functions and are active in the pathogenesis of disorders characterized by chronic inflammation or activation of the coagulation cascade. Signaling through PAR1 and PAR2 shifts the endothelium towards a prothrombotic phenotype, thereby exacerbating the initial pathophysiologic condition. OBJECTIVES: This study aimed to analyze the localization of PARs in the cell membrane and how their compartmentalization affects tissue factor (TF) in human endothelial cells. METHODS: TF expression was determined by quantitative real-time polymerase chain reaction analysis and by activity assays. The interaction of PARs with caveolin was investigated through: (i) caveolin-1 gene knockdown performed by transfection with specific small interfering RNA (siRNA); (ii) caveolin-enriched membrane microdomain disruption; and (iii) coimmunoprecipitation assay. RESULTS: We have shown that PAR1, but not PAR2, is present in endothelial caveolin-enriched membrane microdomains, where it is bound to caveolin-1, and that these structures must be intact if PAR1-induced signaling is to increase TF activity. Cholesterol depletion of endothelial cells by cholesterol-sequestering agents caused the PAR1 to relocate to high-density membranes, and impaired the induction of TF (P < 0.01) without affecting the PAR2-mediated procoagulant effect. In addition, siRNA directed against caveolin-1 inhibited TF activation by PAR1 (P < 0.01 and P < 0.01, respectively). CONCLUSIONS: PAR1 localization in the caveolin-enriched membrane microdomain, bound to caveolin-1, represents a crucial requirement for TF induction in endothelial cells.

A thermodynamic model to predict the thermal response of living beings during pneumoperitoneum procedures.

In this work, hypothermia associated with pneumoperitoneum procedures is studied. A thermodynamic model is developed to allow for the computational simulation of the thermal body response to pneumoperitoneum procedures, which are required by laparoscopic surgery. The numerical results predict the body temperature decay (or loss of energy) in time when the pneumoperitoneum procedures is conducted in patient. The influence of several operating parameters (e.g. inlet air mass flow rate and temperature) on the resulting hypothermia level is analysed. Therefore, the model allows the identification of parameters that have to be controlled to minimize the loss of energy, and consequently, the hypothermia level due to pneumoperitoneum procedures.