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Biological therapies represent the gold-standard treatment of severe forms of plaque psoriasis. However, people living with HIV are often under-treated for psoriasis because very limited data are available on the use of biologics in this population. We report four cases of patients affected by HIV and moderate-to-severe plaque psoriasis, all treated with risankizumab, a monoclonal antibody that selectively targets interleukin-23. After 16 weeks, all patients experienced complete or almost complete skin clearance without any adverse events. Data on the effectiveness and safety of biological therapies in people living with HIV are limited to case reports or small case series, especially for the most recently approved inhibitors of interleukin-23. Our experienced, although limited, supports the role of risankizumab as a safe and effective therapy for psoriasis amongst patients living with HIV.

Aging has been identified as one of the most relevant risk factors for poor outcomes in COVID-19 infection. Since now, different mechanisms responsible for worse outcomes in the elderly have been proposed, which include the remodeling of immune system, the higher prevalence of malnutrition and sarcopenia, the increased burden of multimorbidity, and, to a lesser extent, the direct effects of age on the respiratory system and hormonal profile. It seems that the interplay between all these causes, rather than the individual pathophysiological mechanism, explains the increased severity of the disease with age.

Immune checkpoint inhibitors, such as pembrolizumab and nivolumab, are monoclonal antibodies that block programmed cell death 1 (PD-1) expressed on activated CD8+ T cells and play a crucial role in the treatment of advanced melanoma. With the wide adoption of these therapies, a range of cutaneous adverse effects has been reported, such as flares of plaque psoriasis, but no specific guidelines regarding the treatment are available. We present the case of a 28-year-old male diagnosed with stage IV non-BRAFV600E mutated melanoma in 2014. After the surgery and the failure of ipilimumab and IL-2, he started immunotherapy with pembrolizumab. One month after the start of the therapy, he came to our department showing a severe flare of plaque psoriasis with a body surface area of 40% and a Psoriasis Area and Severity Index (PASI) of 28. Given the severity of the clinical picture and the contraindications to conventional systemic therapy, we decided to start biological treatment with risankizumab, an anti-IL-23 inhibitor. After just the induction phase, he showed almost skin clearance obtaining a reduction of more than 90% of the baseline PASI. Our patient’s rapid response to risankizumab enabled us to continue immunotherapy with pembrolizumab. The recognition of cutaneous signs of toxicity related to such drugs for advanced melanoma is of primary importance to start the correct treatment and continue the immunotherapy when possible.

Cereal flour blending can reduce food insecurity risks, as well as contribute to economic and nutrition goals. Yet, the potential for blending has not been realized, and new products have not become scalable commercial propositions. Numerous experiments have shown the potential to produce acceptable foods derived from blended flours of diverse crops including wheat. An important question is whether the incentives, capacities and needs of farmers, processors and consumers have been considered. We argue that technical solutions must be developed within a specific agroecological, commercial, economic, and political environment. Innovations must address the clearly defined objectives of a wheat flour blending policy, if the potential benefits of blending for addressing food insecurity and economic stress are to be achieved.

Asymmetric catalytic azidation has increased in importance to access enantioenriched nitrogen containing molecules, but methods that employ inexpensive sodium azide remain scarce. This encouraged us to undertake a detailed study on the application of hydrogen bonding phase-transfer catalysis (HB-PTC) to enantioselective azidation with sodium azide. So far, this phase-transfer manifold has been applied exclusively to insoluble metal alkali fluorides for carbon–fluorine bond formation. Herein, we disclose the asymmetric ring opening of mesoaziridinium electrophiles derived from β-chloroamines with sodium azide in the presence of a chiral bisurea catalyst. The structure of novel hydrogen bonded azide complexes was analyzed computationally, in the solid state by X-ray diffraction, and in solution phase by 1H and 14N/15N NMR spectroscopy. With N-isopropylated BINAM-derived bisurea, end-on binding of azide in a tripodal fashion to all three NH bonds is energetically favorable, an arrangement reminiscent of the corresponding dynamically more rigid trifurcated hydrogen-bonded fluoride complex. Computational analysis informs that the most stable transition state leading to the major enantiomer displays attack from the hydrogen-bonded end of the azide anion. All three H-bonds are retained in the transition state; however, as seen in asymmetric HB-PTC fluorination, the H-bond between the nucleophile and the monodentate urea lengthens most noticeably along the reaction coordinate. Kinetic studies corroborate with the turnover rate limiting event resulting in a chiral ion pair containing an aziridinium cation and a catalyst-bound azide anion, along with catalyst inhibition incurred by accumulation of NaCl. This study demonstrates that HB-PTC can serve as an activation mode for inorganic salts other than metal alkali fluorides for applications in asymmetric synthesis.

Homozygous familial hypercholesterolaemia (HoFH) is a rare inherited disorder resulting in extremely elevated low-density lipoprotein cholesterol levels and premature atherosclerotic cardiovascular disease (ASCVD). Current guidance about its management and prognosis stems from small studies, mostly from high-income countries. The objective of this study was to assess the clinical and genetic characteristics, as well as the impact, of current practice on health outcomes of HoFH patients globally.

Fluorine is a key element present in ~35% of agrochemicals and 25% of marketed pharmaceutical drugs. The availability of reliable synthetic protocols to prepare catalysts that allow the efficient incorporation of fluorine in organic molecules is therefore essential for broad applicability. Herein, we report a protocol for the multigram synthesis of two representative enantiopure N-alkyl bis-urea organocatalysts derived from (S)-(–)-1,1′-binaphthyl-2,2′-diamine ((S)-BINAM). These tridentate hydrogen bond donors are highly effective phase-transfer catalysts for solubilizing safe and inexpensive metal alkali fluorides (KF and CsF) in organic solvents for enantioselective nucleophilic fluorinations. The first catalyst, characterized by N-isopropyl substitution, was obtained by using a two-step sequence consisting of reductive amination followed by urea coupling from commercially available starting materials (14 g, 48% yield and 5-d total synthesis time). The second catalyst, featuring N-ethyl alkylation and meta-terphenyl substituents, was accessed via a novel, scalable, convergent route that concluded with the coupling between N-ethylated (S)-BINAM and a preformed isocyanate (52 g and 52% overall yield). On this scale, the synthesis requires ~10 d. This can be reduced to 5 d by performing some steps in parallel. Compared to the previous synthetic route, this protocol avoids the final chromatographic purification and produces the desired catalysts in very high purity and improved yield.

Kernel texture (grain hardness) is a key determinant of wheat quality and utilization. Among soft wheats, softer kernels possess some advantages in processing and end‐product quality. Here, an extra soft kernel phenotype was identified and characterized in soft kernel durum wheat. A RIL population was developed by crossing “Soft Strongfield” with Kamut Khorasan wheat. Progeny kernels were selected for chalky appearance in the F2, F3,and F4. Kernel texture phenotypes measured by the SKCS in the F5, F6,and F7were all less than 20 and ranged down to −12.6. Among F5:7full‐sib RILs, there was indication of a single segregating genetic locus that was associated with a difference of about 7 SKCS hardness index (HI) units. Single marker–trait association and composite interval mapping both identified significant QTLs on chromosomes 4B and 1B with phenotypic effects of about 5 and 4 HI, respectively. QTL detected in the present study may be used to reduce kernel texture in soft durum to below zero, creating “extra soft” durum. These QTLs should also be available to be used in hard kernel durum to reduce kernel texture and may be transferrable to hexaploid wheat. The creation of soft kernel durum wheat expanded the processing, end‐product quality, and utilization of durum wheat. A further reduction in kernel softness may expand further durum wheat's importance as a global food.

Utilization of durum wheat (Triticum turgidumsubsp. durum) can be enhanced by increasing grain and flour protein content. One strategy to increase protein content is by introducing the functional Gpc‐B1allele from wild emmer (Triticum turgidumsubsp. dicoccoides). Introduction of the functional Gpc‐B1allele into soft kernel durum increased grain and flour protein by 17 g/kg, increased dough strength as evidenced by SDS sedimentation volume and Mixograph dough mixing parameters, and increased straight‐dough pan bread volume. When grown under arid conditions, high protein (151 g/kg) samples had decreased loaf volumes indicative of inelastic doughs. The functional Gpc‐B1allele was associated with decreased test weight, a small increase in SKCS hardness, and a modest increase in flour ash; otherwise, milling performance was not affected. Introgression of the Gpc‐B1functional allele from dicoccoides into durum wheat can improve dough strength and breadmaking quality. The effect tends to be consistent over environments but overall, Gpc‐B1made only a modest improvement in durum wheat breadmaking quality. Further studies with concomitant selection at other loci are needed to see the effects of Gpc‐B1among elite germplasm. Durum wheat production and consumption will increase as bread quality improves. The functional Gpc‐B1allele contributed to improved breadmaking quality. The present report is the first to examine the effect of this allele on breadmaking in durum wheat.

Growth hormone (GH) deficiency (GHD) in children is defined as impaired production of GH by the pituitary gland that results in growth failure. This disease might be congenital or acquired, and occurs in isolation or in the setting of multiple pituitary hormone deficiency. Isolated GHD has an estimated prevalence of 1 patient per 4000–10,000 live births and can be due to multiple causes, some of which are yet to be determined. Establishing the correct diagnosis remains key in children with short stature, as initiating treatment with recombinant human GH can help them attain their genetically determined adult height. During the past two decades, our understanding of the benefits of continuing GH therapy throughout the transition period from childhood to adulthood has increased. Improvements in transitional care will help alleviate the consequent physical and psychological problems that can arise from adult GHD, although the consequences of a lack of hormone replacement are less severe in adults than in children. In this Review, we discuss the differential diagnosis in children with GHD, including details of clinical presentation, neuroimaging and genetic testing. Furthermore, we highlight advances and issues in the management of GHD, including details of transitional care.

Selection for high breadmaking quality is mostly conducted using tests employing refined flour and added salt which may not be adequate for selecting lines with high wholemeal or low‐sodium breadmaking potential. The objective of this study was thus to evaluate the suitability of the current breadmaking quality tests for the selection of wheat lines intended for high‐quality healthier breads. A set of common wheat lines was analyzed for its breadmaking quality using three bread formulations: refined flour and 1% salt (RF); refined flour and no added salt (RNa); and reconstituted flour and 1% salt (WM). The RNa bread quality was accurately predicted using the RF formulation (r> 0.90) and was highly correlated with protein content and dough properties. Differently, the WM breadmaking quality was less predictable using the RF formulation (r= 0.64) and was highly correlated with grain hardness and dough properties but less with protein content. The current methods used for evaluating wheat breadmaking quality are effective to predict low‐sodium bread quality but not wholemeal bread quality. Results of this study highlighted the need of using wholemeal rather than refined flour for the selection of wheat lines with high wholemeal breadmaking potential and suggested that traits such as grain hardness and SDS‐sedimentation volume could be useful to predict high wholemeal breadmaking quality.

Utilization of durum wheat (Triticum turgidumssp. durum) is limited by its weak gluten and poor bread‐making quality. One strategy to improve gluten strength and bread‐making quality is by introducing the Glu‐D1alleles from bread wheat. Introduction of Glu‐D1alleles Glu‐D1aand Glu‐D1d, corresponding to the High Molecular Weight Glutenin subunits Dx2 + Dy12 and Dx5 + Dy10, respectively, increased dough strength as evidenced by SDS sedimentation volume, Lactic acid Solvent Retention Capacity, and Mixograph dough mixing parameters. The Glu‐D1dDx5 + Dy10 allele was “stronger” than the Glu‐D1aDx2 + Dy12 allele. However, whereas Dx2 + Dy12 improved straight‐dough pan bread volume, Dx5 + Dy10 did not. This latter result was ascribed to the overly strong, inextensible gluten contributed by the Glu‐D1dDx5 + Dy10 allele. Whereas both Glu‐D1alleles increased dough strength, Dx2 + Dy12 improved bread loaf volume but lines with Dx5 + Dy10 produced doughs that were too strong to realize an increase in loaf volume. The results confirm the notion that large bread loaf volume is achieved with a balance of dough extensibility and elasticity. Durum wheat production and consumption will increase as bread quality improves. The Glu‐D1high molecular weight glutenin proteins will likely play a role in improving bread‐making ability. The two major Glu‐D1alleles are accessible via durum wheat translocation lines.

AbstractThe Pecorino Romano P.D.O. is the main sheep cheese produced in Italy and the first one among the sheep cheeses, in terms of quality and value, exported from the European Union. About half of the sheep milk produced in Italy is processed into this type of cheese by 36 dairies belonging to the Pecorino Romano Consortium. Eight million records of biweekly analyses of milk collected within a 5-year period from farms delivering their milk to the aforementioned consortium were analysed in this work. Monthly evolution curves were plotted for fat, protein, lactose, pH, NaCl, SCC, bacterial load and principal fatty acids (FAs). Due to the seasonal production systems of Sardinian sheep, monthly evolution of milk fat and protein contents and cheese yield are directly linked to the lactation curve pattern and the pastures quantity and quality. Also, the FA profile of milk is affected by grass availability and quality in both early and mid-lactation, whereas it is influenced by the energy balance of ewes in late lactation. Cheese yield equation was computed based on fat and protein contents and considering the variability among dairies in technological processes used in transforming Sarda sheep milk to Pecorino Romano P.D.O. These data could be a relevant basis to set-up future grids of milk payments based on quality standards. Moreover, they could be useful to formulate administrative policies on the dairy sector with the prospective to improve milk quality of Sardinian sheep destined to the Pecorino Romano production.HighlightsThe yield of Pecorino Romano P.D.O. (PR) is linked to fat and protein content of milk and it depends on the industrial processes adopted by each dairyMonthly variation of milk composition is of high interest for the PR producers to program the output of their milk-processing plansThe content and monthly evolutions of principal fatty acids are important to determine the nutritional and technological properties of PR

A number of studies have evaluated the role of IGF1 measurement in the diagnosis of growth hormone deficiency (GHD). This study aimed to evaluate the accuracy and the best cut-off of IGF1 SDS in the diagnosis of GHD in a large cohort of short children and adolescents. One-hundred and forty-two children and adolescents with GHD ((63 organic/genetic (OGHD), 79 idiopathic (IGHD)) and 658 short non-GHD children (median age 10.4 years) were included in the analysis. The two groups were subdivided according to age (G1 <6, G2 6 <9, G3 9 <12, G4 ≥12) and to pubertal status. Serum IGFI was measured by the same chemiluminescence assay in all samples and expressed as age- and sex-based SDS. Receiver operating characteristic (ROC) analysis was used to evaluate the optimal IGF1 SDS cut-off and the diagnostic accuracy. Median IGF1 SDS was significantly lower in the GHD than in non-GHD patients. The area under the curve (AUC) was 0.69, with the best IGF1 cut-off of −1.5 SDS (sensitivity 67.61%, specificity 62.62%). The AUC was 0.75 for OGHD and 0.63 for IGHD. The accuracy was better in the pubertal (AUC = 0.81) than the prepubertal group (AUC = 0.64). In our cohort, IGF1 measurement has poor accuracy in discriminating GHD from non-GHD. Our findings confirm and reinforce the belief that IGF1 values should not be used alone in the diagnosis of GHD but should be interpreted in combination with other clinical and biochemical parameters.

Ammonium salts are used as phase-transfer catalysts for fluorination with alkali metal fluorides. We now demonstrate that these organic salts, specifically azetidinium triflates, are suitable substrates for enantioselective ring opening with CsF and a chiral bis-urea catalyst. This process, which highlights the ability of hydrogen bonding phase-transfer catalysts to couple two ionic reactants, affords enantioenriched γ-fluoroamines in high yields. Mechanistic studies underline the role of the catalyst for phase-transfer, and computed transition state structures account for the enantioconvergence observed for mixtures of achiral azetidinium diastereomers. The N-substituents in the electrophile influence the reactivity, but the configuration at nitrogen is unimportant for the enantioselectivity.

Background: Coxsackievirus infections are associated with cases of aseptic meningitis, encephalitis, myocarditis, and some chronic disease. Methods: A series of benzo[d][1,2,3]triazol-1(2)-yl derivatives (here named benzotriazol-1(2)-yl) (4a-i, 5a-h, 6a-e, g, i, j and 7a-f, h-j) were designed, synthesized and in vitro evaluated for cytotoxicity and antiviral activity against two important human enteroviruses (HEVs) members of the Picornaviridae family [Coxsackievirus B 5 (CVB-5) and Poliovirus 1 (Sb-1)]. Results: Compounds 4c (CC50 >100 μM; EC50 = 9 μM), 5g (CC50 >100 μM; EC50 = 8 μM), and 6a (CC50 >100 μM; EC50 = 10 μM) were found active against CVB-5. With the aim of evaluating the selectivity of action of this class of compounds, a wide spectrum of RNA (positive- and negativesense), double-stranded (dsRNA) or DNA viruses were also assayed. For none of them, significant antiviral activity was determined. Conclusion: These results point towards a selective activity against CVB-5, an important human pathogen that causes both acute and chronic diseases in infants, young children, and immunocompromised patients.

The aminoacyl-tRNA synthetases are an essential and universally distributed family of enzymes that plays a critical role in protein synthesis, pairing tRNAs with their cognate amino acids for decoding mRNAs according to the genetic code. Synthetases help to ensure accurate translation of the genetic code by using both highly accurate cognate substrate recognition and stringent proofreading of noncognate products. While alterations in the quality control mechanisms of synthetases are generally detrimental to cellular viability, recent studies suggest that in some instances such changes facilitate adaption to stress conditions. Beyond their central role in translation, synthetases are also emerging as key players in an increasing number of other cellular processes, with far-reaching consequences in health and disease. The biochemical versatility of the synthetases has also proven pivotal in efforts to expand the genetic code, further emphasizing the wide-ranging roles of the aminoacyl-tRNA synthetase family in synthetic and natural biology.

Hydrogen-bonding interactions have been explored in catalysis, enabling complex chemical reactions. Recently, enantioselective nucleophilic fluorination with metal alkali fluoride has been accomplished with BINAM-derived bisurea catalysts, presenting up to four NH hydrogen-bond donors (HBDs) for fluoride. These catalysts bring insoluble CsF and KF into solution, control fluoride nucleophilicity, and provide a chiral microenvironment for enantioselective fluoride delivery to the electrophile. These attributes encouraged a 1H/19F NMR study to gain information on hydrogen-bonding networks with fluoride in solution, as well as how these arrangements impact the efficiency of catalytic nucleophilic fluorination. Herein, NMR experiments enabled the determination of the number and magnitude of HB contacts to fluoride for thirteen bisurea catalysts. These data supplemented by diagnostic coupling constants 1hJNH···F–give insight into how multiple H bonds to fluoride influence reaction performance. In dichloromethane (DCM-d2), nonalkylated BINAM-derived bisurea catalyst engages two of its four NH groups in hydrogen bonding with fluoride, an arrangement that allows effective phase-transfer capability but low control over enantioselectivity for fluoride delivery. The more efficient N-alkylated BINAM-derived bisurea catalysts undergo urea isomerization upon fluoride binding and form dynamically rigid trifurcated hydrogen-bonded fluoride complexes that are structurally similar to their conformation in the solid state. Insight into how the countercation influences fluoride complexation is provided based on NMR data characterizing the species formed in DCM-d2when reacting a bisurea catalyst with tetra-n-butylammonium fluoride (TBAF) or CsF. Structure–activity analysis reveals that the three hydrogen-bond contacts with fluoride are not equal in terms of their contribution to catalyst efficacy, suggesting that tuning individual electronic environment is a viable approach to control phase-transfer ability and enantioselectivity.

The photoinduced, multicomponent reaction of α-diazoketones, silanols, and isocyanides affords α-silyloxy acrylamides, formally derived from α-keto amides. The presence of a secondary amido group makes classic preparative methods for silyl enol ethers unfeasible in this case, while the mild conditions required by this photochemical approach allow their synthesis in good yields; moreover, the general structure can be easily modified by varying each component of the multicomponent reaction. Fine-tuning of the reaction conditions (i.e., solvents, radiation, additives) can be exploited to obtain complete Zselectivity. The reactivity of this overlooked class of silyl enol ethers has been investigated, and features that could pave the way to new applications have been found.

The use of plant growth-promoting bacteria as bioinoculants is a powerful tool to increase crop yield and quality and to improve nitrogen use efficiency (NUE) from fertilizers in plants. This study aimed to bioprospecting a native bacterial consortium (Bacillus cabrialesiisubsp. cabrialesiiTE3T, Priestia megateriumTRQ8, and Bacillus paralicheniformisTRQ65), through bioinformatic analysis, and to quantify the impact of its inoculation on NUE (measured through 15N-isotopic techniques), grain yield, and grain quality of durum wheat variety CIRNO C2008 grown under three doses of urea (0, 120, and 240 kg N ha−1) during two consecutive agricultural cycles in the Yaqui Valley, Mexico. The inoculation of the bacterial consortium (BC) to the wheat crop, at a total N concentration of 123–225 kg N ha−1increased crop productivity and maintained grain quality, resulting in a yield increase of 1.1 ton ha−1(6.0 vs. 7.1 ton ha−1, 0 kg N ha−1added, 123 kg N ha−1in the soil) and of 2.0 ton ha−1(5.9 vs. 7.9 ton ha−1, 120 kg N ha−1added, 104 kg N ha−1in the soil) compared to the uninoculated controls at the same doses of N. The genomic bioinformatic analysis of the studied strains showed a great number of biofertilization-related genes regarding N and Fe acquisition, P assimilation, CO2fixation, Fe, P, and K solubilization, with important roles in agroecosystems, as well as genes related to the production of siderophores and stress response. A positive effect of the BC on NUE at the studied initial N content (123 and 104 kg N ha−1) was not observed. Nevertheless, increases of 14 % and 12.5 % on NUE (whole plant) were observed when 120 kg N ha−1was applied compared to when wheat was fully fertilized (240 kg N ha−1). This work represents a link between bioinformatic approaches of a native bacterial inoculant and the quantification of its impact on durum wheat.

Potassium fluoride (KF) is an ideal reagent for fluorination because it is safe, easy to handle and low-cost. However, poor solubility in organic solvents coupled with limited strategies to control its reactivity has discouraged its use for asymmetric C–F bond formation. Here, we demonstrate that hydrogen bonding phase-transfer catalysis with KF provides access to valuable β-fluoroamines in high yields and enantioselectivities. This methodology employs a chiral N-ethyl bis-urea catalyst that brings solid KF into solution as a tricoordinated urea-fluoride complex. This operationally simple reaction affords enantioenriched fluoro-diphenidine (up to 50 g scale) using 0.5 mol % of recoverable bis-urea catalyst.

Background:Acute lymphoblastic leukemia (ALL) is a cancer of lymphoid cells and organs that requires immunochemotherapy combined with hematopoietic stem cell transplantation (HSCT) according to minimal residual disease. Recently, cell therapy and targeted drugs have been added to the treatment algorithm. Supportive care, and local treatment such as intrathecal chemotherapy are also recommended due to complications related to disease and therapy.

Background: Acute myeloid leukemia (AML) is a cancer of the blood and bone marrow cells that requires intensive chemotherapy combined with hematopoietic stem cell transplantation in fit patients and less intensive treatment in elderly patients. Supportive care is also advised due to disease and therapy-related cytopenia that can lead to life-threatening systemic infections, bleeding, and hypoxemia.

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ABSTRACTThe aminoacylation status of the cellular tRNA pool regulates both general amino acid control (GAAC) and target of rapamycin (TOR) stress response pathways in yeast. Consequently, fidelity of translation at the level of aminoacyl-tRNA synthesis plays a central role in determining accuracy and sensitivity of stress responses. To investigate effects of translational quality control (QC) on cell physiology under stress conditions, phenotypic microarray analyses were used to identify changes in QC deficient cells. Nitrogen source growth assays showed QC deficient yeast grew differently compared to WT. The QC deficient strain was more tolerant to caffeine treatment than wild type through altered interactions with the TOR and GAAC pathways. Increased caffeine tolerance of the QC deficient strain was consistent with the observation that the activity of Gln3p, a transcription factor controlled by the TOR pathway, is decreased in the QC deficient strain compared to WT. GCN4 translation, which is typically repressed in the absence of nutritional stress, was enhanced in the QC deficient strain through TOR inhibition. QC did not impact cell cycle regulation; however, the chronological lifespan of QC deficient yeast strains decreased compared to wild type, likely due to translational errors and alteration of the TOR-associated regulon. These findings support the idea that changes in translational fidelity provide a mechanism of cellular adaptation by modulating TOR activity. This, in turn, supports a central role for aminoacyl-tRNA synthesis QC in the integrated stress response by maintaining the proper aa-tRNA pools necessary to coordinate the GAAC and TOR.

Improved treatments are needed for hemophilia A and B, bleeding disorders affecting 400 000 people worldwide. We investigated whether targeting protein S could promote hemostasis in hemophilia by rebalancing coagulation. Protein S (PS) is an anticoagulant acting as cofactor for activated protein C and tissue factor pathway inhibitor (TFPI). This dual role makes PS a key regulator of thrombin generation. Here, we report that targeting PS rebalances coagulation in hemophilia. PS gene targeting in hemophilic mice protected them against bleeding, especially when intra-articular. Mechanistically, these mice displayed increased thrombin generation, resistance to activated protein C and TFPI, and improved fibrin network. Blocking PS in plasma of hemophilia patients normalized in vitro thrombin generation. Both PS and TFPIα were detected in hemophilic mice joints. PS and TFPI expression was stronger in the joints of hemophilia A patients than in those of hemophilia B patients when receiving on-demand therapy, for example, during a bleeding episode. In contrast, PS and TFPI expression was decreased in hemophilia A patients receiving prophylaxis with coagulation factor concentrates, comparable to osteoarthritis patients. These results establish PS inhibition as both controller of coagulation and potential therapeutic target in hemophilia. The murine PS silencing RNA approach that we successfully used in hemophilic mice might constitute a new therapeutic concept for hemophilic patients.

Improved treatments are needed for hemophilia A and B, bleeding disorders affecting 400 000 people worldwide. We investigated whether targeting protein S could promote hemostasis in hemophilia by rebalancing coagulation. Protein S (PS) is an anticoagulant acting as cofactor for activated protein C and tissue factor pathway inhibitor (TFPI). This dual role makes PS a key regulator of thrombin generation. Here, we report that targeting PS rebalances coagulation in hemophilia. PS gene targeting in hemophilic mice protected them against bleeding, especially when intra-articular. Mechanistically, these mice displayed increased thrombin generation, resistance to activated protein C and TFPI, and improved fibrin network. Blocking PS in plasma of hemophilia patients normalized in vitro thrombin generation. Both PS and TFPIα were detected in hemophilic mice joints. PS and TFPI expression was stronger in the joints of hemophilia A patients than in those of hemophilia B patients when receiving on-demand therapy, for example, during a bleeding episode. In contrast, PS and TFPI expression was decreased in hemophilia A patients receiving prophylaxis with coagulation factor concentrates, comparable to osteoarthritis patients. These results establish PS inhibition as both controller of coagulation and potential therapeutic target in hemophilia. The murine PS silencing RNA approach that we successfully used in hemophilic mice might constitute a new therapeutic concept for hemophilic patients.

La Sindrome di Down ha un’incidenza di 1/400–1500 nati vivi ed è la causa più frequente di disabilità intellettiva di origine genetica. È caratterizzata da un insieme di manifestazioni fenotipiche variabili riscontrabili sin dalla nascita, legate alla presenza, parziale o completa, di un cromosoma 21 sovrannumerario. Oltre alle caratteristiche fisiche che la contraddistinguono, la Sindrome di Down può presentare complicanze a livello sistemico. Le complicanze endocrine più frequenti della sindrome di Down sono a carico della tiroide, dell’osso, del sistema metabolico, delle gonadi e del pancreas.

Systemic sclerosis (SSc) is characterized by endothelial cell (EC) apoptosis, impaired angiogenesis and peripheral microvasculopathy. Soluble α-Klotho (sKl) is a pleiotropic molecule with multiple effects on ECs, including antioxidant and vasculoprotective activities. On the EC surface, sKl interacts with vascular endothelial growth factor (VEGF) receptor-2 (VEGFR-2) and transient receptor potential canonical-1 (TRPC-1) cation channel to control EC homeostasis. Here, we investigated whether sKl might act as a protective factor to improve angiogenesis in dermal microvascular endothelial cells (MVECs) from SSc patients (SSc-MVECs). Wound healing assay was performed on healthy dermal MVECs (H-MVECs) challenged with sera from healthy controls or SSc patients with or without the addition of sKl. Capillary morphogenesis on Matrigel was assessed in H-MVECs and SSc-MVECs at basal conditions and treated with sKl, as well as in H-MVECs challenged with healthy or SSc sera in presence or absence of sKl. The expression of α-Klotho, VEGF165b, VEGFR-2, TRPC-1, Ki67 and active caspase-3 in H-MVECs and SSc-MVECs was investigated by western blotting. Immunostaining for α-Klotho was performed in H-MVECs and SSc-MVECs, and in healthy and SSc skin sections. Treatment with sKl effectively counteracted the inihibitory effects of SSc sera on wound healing ability and angiogenic performance of H-MVECs. The addition of sKl significantly improved angiogenesis and maintained over time capillary-like tube formation in vitro by SSc-MVECs. Stimulation of SSc-MVECs with sKl resulted in the upregulation of the proliferation marker Ki67 in parallel with the downregulation of proapoptotic active caspase-3. The expression of α-Klotho was significantly lower in SSc-MVECs than in H-MVECs. The expression of TRPC-1 was also significantly decreased, while that of VEGFR-2 and VEGF165b was significantly increased, in SSc-MVECs compared with H-MVECs. Challenge with sKl either significantly increased TRPC-1 or decreased VEGF165b in SSc-MVECs. Ex vivo analyses revealed that α-Klotho immunostaining was almost absent in the dermal microvascular network of SSc skin compared with control skin. Our findings provide the first evidence that α-Klotho is significantly decreased in the microvasculature in SSc skin and that sKl administration may effectively improve SSc-MVEC functions in vitro by acting as a powerful proangiogenic factor.

The low-molecular-weight glutenin subunits (LMW-GSs) are a class of wheat seed storage proteins encoded by a multigene family located at the Glu-3loci that influences wheat end-use quality. Owing to ambiguities in the LMW-GS allele nomenclature and to the complexity of the Glu-3loci organization, a clear relationship between LMW-GS alleles and wheat end-use quality has not been adequately determined. In the present study, four sets of elite common wheat varieties were analyzed for their LMW-GS genic profile, along with their dough rheology and end-product baking properties. Among these varieties, variation at the Glu-A3locus had a major impact on the analyzed dough rheology parameters, followed by the Glu-B3and Glu-D3loci. Also, the genes located at the linkage groups Glu-A3-3, Glu-B3-3, and Glu-D3-5 were more highly associated with dough strength, mixing, and extensibility properties. Results obtained in this study clearly indicate that there are specific LMW-GS haplotypes that are more highly associated than others to variation in dough rheology.

Durum wheat (Triticum turgidumssp. durum) is considered unsuitable for the majority of commercial bread production because of its weaker gluten strength, combined with a larger flour particle size and higher level of starch damage after milling. Recently, a new durum cultivar with soft kernel texture, Soft Svevo, was developed by the Ph1b‐mediated homoeologous transfer of the Puroindolinegenes at the Hardness(Ha) locus from the D genome of T. aestivumL. The objective of this research was to evaluate the dough strength and pan bread‐making potential of soft‐kernel durum germplasm developed from crossing Soft Svevo to selected entries of the International Maize and Wheat Improvement Center (CIMMYT) 44th International Durum Yield Nursery. Forty‐six F2:5soft durum full and half‐sib lines were grown in replicated plots in two locations. Grain samples were evaluated for flour protein quality, dough mixing, and strength characteristics and bread baking. Significant differences (p< 0.05) were detected among lines for flour sodium dodecylsulfate (SDS) sedimentation volume (3.2–12.6 mL g−1), solvent retention capacity lactic acid (63.2–112.6 g 100 g−1), Mixograph water absorption (59.8–64.5 g 100 g−1), peak height (37.1–52.8), and peak width (55.4–125.2), and loaf volume (629–864 cm3).These results indicate that the bread‐making potential of soft durum can be improved if hard durum cultivars with favorable alleles for protein quality, dough strength, and bread making are chosen as crossing parents with Soft Svevo and, by extension, other soft‐kernel durum wheat germplasm.

Spelt (Triticum aestivum ssp. spelta) is part of the so-called ancient wheats. These types of wheats are experiencing a revival as they have been proposed to be healthier than conventional wheat. However, the given healthier condition of spelt is not substantiated by solid scientific evidence. The objective of this study was to analyze the genetic variability for several grain components, related to nutritional quality (arabinoxylans, micronutrients, phytic acid) in a set of spelt and common wheat genotypes to determinate if spelt is potentially healthier than common wheat. The results obtained indicated that within the compared species, there is a significant variation in the nutritional compounds, and it is not truthful and accurate to state that one species is healthier than the other. Within both groups, genotypes showing outstanding values for some traits were detected, which could be used in breeding programs to develop new wheat cultivars with good agronomic performance and nutritional quality.

Structural and functional diversity among the aminoacyl-tRNA synthetases prevent infiltration of the genetic code by noncognate amino acids. To explore whether these same features distinguish the synthetases as potential sources of resistance against antibiotic amino acid analogues, we investigated bacterial growth inhibition by S-(2-aminoethyl)- l-cysteine (AEC). Wild-type lysyl-tRNA synthetase (LysRS) and a series of active site variants were screened for their ability to restore growth of an Escherichia coliLysRS null strain at increasing concentrations of AEC. While wild-type E. coligrowth is completely inhibited at 5 µM AEC, two LysRS variants, Y280F and F426W, provided substantial resistance and allowed E. colito grow in the presence of up to 1 mM AEC. Elevated resistance did not reflect changes in the kinetics of amino acid activation or tRNA Lysaminoacylation, which showed at best 4−6-fold improvements, but instead correlated with the binding affinity for AEC, which was decreased ∼50-fold in the LysRS variants. In addition to changes in LysRS, AEC resistance has also been attributed to mutations in the L box riboswitch, which regulates expression of the lysCgene, encoding aspartokinase. The Y280F and F426W LysRS mutants contained wild-type L box riboswitches that responded normally to AEC in vitro, indicating that LysRS is the primary cellular target of this antibiotic. These findings suggest that the AEC resistance conferred by L box mutations is an indirect effect resulting from derepression of lysCexpression and increased cellular pools of lysine, which results in more effective competition with AEC for binding to LysRS.

The vascular and nervous systems have several anatomic and molecular mechanism similarities. Emerging evidence suggests that proteins involved in transmitting axonal guidance cues, including members of class III semaphorin (Sema3) family, play a critical role in blood vessel guidance during physiological and pathological vascular development. Sema3E is a natural antiangiogenic molecule that causes filopodial retraction in endothelial cells, inhibiting cell adhesion by disrupting integrin-mediated adhesive structures. The aim of the present study was to investigate whether in systemic sclerosis (SSc) Plexin-D1/Sema3E axis could be involved in the dysregulation of vascular tone control and angiogenesis. Sema3E levels were measured by quantitative colorimetric sandwich ELISA in serum samples from 48 SSc patients, 45 subjects with primary Raynaud's phenomenon (pRP) and 48 age-matched and sex-matched healthy controls. Immunofluorescence staining on skin sections from 14 SSc patients and 12 healthy subjects was performed to evaluate Sema3E and Plexin-D1 expression. Western blotting was used to assess Plexin-D1/Sema3E axis in human SSc and healthy dermal microvascular endothelial cells (SSc-MVECs and H-MVECs, respectively) at basal condition and after stimulation with recombinant human vascular endothelial growth factor (VEGF), SSc and healthy sera. Capillary morphogenesis on Matrigel was performed on H-MVECs treated with healthy, pRP or SSc sera in the presence of Sema3E and Plexin-D1 soluble peptides. Serum Sema3E levels were significantly higher both in pRP subjects and SSc patients than in controls. In SSc, Sema3E levels were significantly increased in patients with early nailfold videocapillaroscopy (NVC) pattern compared to active/late patterns and pRP, and in patients without digital ulcers versusthose with ulcers. In SSc skin, Sema3E expression was strongly increased in the microvascular endothelium. Cultured SSc-MVECs showed higher levels of phosphorylated Plexin-D1 and Sema3E expression than H-MVECs, and stimulation with SSc sera increased phosphorylated Plexin-D1 and Sema3E in H-MVECs. The addition of Sema3E-binding Plexin-D1 soluble peptide significantly attenuated the antiangiogenic effect of SSc sera on H-MVECs. Our findings suggest that Plexin-D1/Sema3E axis is triggered in SSc endothelium and may have a role in the dysregulation of angiogenesis and vascular tone control by inducing neuro-vascular mechanism alterations clinically evident in particular in the early disease phases.

Sequence-specific interactions between aminoacyl-tRNA synthetases and their cognate tRNAs ensure both accurate RNA recognition and the efficient catalysis of aminoacylation. The effects of tRNA(Trp)variants on the aminoacylation reaction catalyzed by wild-type Escherichia coli tryptophanyl-tRNA synthe-tase (TrpRS) have now been investigated by stopped-flow fluorimetry, which allowed a pre-steady-state analysis to be undertaken. This showed that tRNA(Trp)identity has some effect on the ability of tRNA to bind the reaction intermediate TrpRS-tryptophanyl-adenylate, but predominantly affects the rate at which trypto-phan is transferred from TrpRS-tryptophanyl adenylate to tRNA. Use of the binding ( K (tRNA)) and rate constants ( k (4)) to determine the energetic levels of the various species in the aminoacylation reaction showed a difference of approximately 2 kcal mol(-1)in the barrier to transition state formation compared to wild-type for both tRNA(Trp)A-->C73 and. These results directly show that tRNA identity contributes to the degree of complementarity to the transition state for tRNA charging in the active site of an aminoacyl-tRNA synthetase:aminoacyl-adenylate:tRNA complex.

Context:Although balance is generally recognized to be an important feature in ensuring good performance in soccer, its link with functional performance remains mostly unexplored, especially in young athletes.Objective:To investigate changes in balance induced by fatigue for unipedal and bipedal static stances in young soccer players.Design:Crossover study.Setting:Biomechanics laboratory and outdoor soccer field.Patients or Other Participants:Twenty-one male soccer players (age = 14.5 ± 0.2 years, height = 164.5 ± 5.6 cm, mass = 56.8 ± 6.8 kg).Intervention(s):Static balance was assessed with postural-sway analysis in unipedal and bipedal upright stance before and after a fatigue protocol consisting of a repeated sprint ability (RSA) test (2 × 15-m shuttle sprint interspersed with 20 seconds of passive recovery, repeated 6 times).Main Outcome Measure(s):On the basis of the center-of-pressure (COP) time series acquired during the experimental tests, we measured sway area, COP path length, and COP maximum displacement and velocity in the anteroposterior and mediolateral directions.Results:Fatigue increased all sway values in bipedal stance and all values except COP velocity in the mediolateral direction in unipedal stance. Fatigue index (calculated on the basis of RSA performance) was positively correlated with fatigue/rest sway ratio for COP path length and COP velocity in the anteroposterior and mediolateral directions for nondominant single-legged stance.Conclusions:Fatigued players exhibited reduced performance of the postural-control system. Participants with better performance in the RSA test appeared less affected by balance impairment, especially in single-legged stance.

Listeria monocytogenesis an intracellular, foodborne gastrointestinal pathogen that is primarily responsible for causing listeriosis or food poisoning in otherwise healthy individuals. Infections that arise during pregnancy or within immune compromised individuals are much more serious resulting in the risk of fetal termination or fetal fatality postpartum in the former and septicemia or meningitis with a 20% fatality rate in the latter. While the roles of internalin proteins and listeriolysin-O in the infection process are well characterized, the specific roles of lysine-modified phospholipids in the membrane of L. monocytogenesare not. Investigation into the lipid bilayer composition of L. monocytogenesindicated that the overall proportions of lipids, including lysylcardiolipin and lysylphosphatidylglycerol (LysPG), vary with growth temperature and growth phase. In addition, we demonstrate that LysPG formation is essential for L. monocytogenessurvival in the presence of increased osmolytic stress but has no effect on bacterial adherence, invasion or survival in the presence of physiologically relevant concentrations of human neutrophil peptide (HNP-1). In the absence of LysPG synthesis, L. monocytogenesunexpectedly retained flagellum-mediated motility at 37 °C. Taken together, these findings show that LysPG formation in L. monocytogeneshas broader functions in virulence and survival beyond its known role in the modification of membrane potential previously observed in other bacteria.

In obese subjects it has been shown that cortisol (F) contributes to the reduction in insulin sensitivity, suggesting a role in the development of the metabolic syndrome (MS). The aim of this retrospective study was to evaluate the relationship between F and components of MS in 1,027 obese children and adolescents. Waist circumference, systolic and diastolic blood pressure (SP, DP), F, serum glucose (Glyc), cholesterol HDL, triglycerides and homeostatic model assessment (HOMA index) were evaluated in all subjects. MS was defined according to the International Diabetes Federation criteria. Accordingly, patients were subdivided into three age groups: 6–10, 10–16 and >16 years. In univariate regression analysis, F was correlated with Glyc, SP and HOMA in groups 1 and 2, with DP in Group 2. In multivariate regression analysis including age, sex, puberty, BMI-SDS and F as independent variables and one of the component of the MS as the dependent variable, F was a weak predictor of the variability when DP and Glyc were introduced as dependent variables in Group 2 and when SP was introduced as dependent variable both in groups 1 and 2. When patients were subdivided into subgroups according to the IDF criteria, in Group 2 patients with one or more components of the MS had higher F concentrations. In this cohort of obese children and adolescents, F was weakly associated with components of the MS. These findings do not support a major role for F in the development of MS.

AbstractAim:We aimed to study the influence of the fat mass and obesity-associated (FTO) gene on eating behavior in 412 obese Sardinian children and adolescents. Genome-wide association studies (GWAS) have identified several susceptibility loci for obesity. Among these, the polymorphisms in the intron 1 of the FTOgene has been found associated to weight gain and obesity in various populations.Methods: All obese patients were genotyped for the FTOsingle nucleotide polimorphysm (SNP) rs9939609. In all subjects we evaluated eating behavior using the Child Eating Behaviour Questionnaire (CEBQ).Results:We found no differences in eating behavior according to the genotype, either in the entire cohort, or when subjects were subdivided into four different age groups.Conclusions: FTOgenotype is associated with body mass index but does not influence eating behavior in a selected cohort of obese children from the isolated genetic population of Sardinia.

Objective.Hemarthrosis triggers hemophilic arthropathy, involving the target joints. The histopathogenesis of blood-induced joint damage remains unclear. The triad of receptor activator of nuclear factor-κB (RANK), RANK ligand (RANKL), and osteoprotegerin (OPG; RANK-RANKL-OPG) controls bone turnover. Our aim was to evaluate RANK-RANKL-OPG expression in the synovium of hemophilic patients with severe arthropathy.Methods.Synovial biopsies were obtained from 18 patients with hemophilic arthropathy and 16 with osteoarthritis (OA) who were undergoing total knee replacement and synovectomy. The severity of hemophilic arthropathy was evaluated according to ultrasonography score, the World Federation of Hemophilia (WFH) orthopedic joint scale, and the radiographic Pettersson score. RANK-RANKL-OPG expression was examined by immunohistochemistry and Western blotting. Serum levels of soluble RANKL (sRANKL) and OPG from an extended group of 67 patients with hemophilic arthropathy and 30 healthy controls were measured by ELISA.Results.The mean ultrasonography, WFH orthopedic joint scale, and Pettersson scores in patients with hemophilic arthropathy indicated severe arthropathy. A decreased expression of OPG was found in hemophilic arthropathy synovium compared with patients with OA. RANK and RANKL immunopositivity was strong in the lining and sublining layers in hemophilic arthropathy synovial tissue. Western blotting confirmed the immunohistological findings. Serum levels of sRANKL and OPG in patients with hemophilia were lower than in healthy controls.Conclusion.In hemophilic arthropathy, the synovium highly expressed RANK and RANKL, whereas OPG immunopositivity decreased, suggesting an osteoclastic activation. Low tissue expression of OPG paralleled the serum levels of this protein and the severity of hemophilic arthropathy assessed by ultrasonography, Pettersson, and WFH orthopedic joint scale scores.

In 2008 we published the first set of guidelines for standardizing research in autophagy. Since then, research on this topic has continued to accelerate, and many new scientists have entered the field. Our knowledge base and relevant new technologies have also been expanding. Accordingly, it is important to update these guidelines for monitoring autophagy in different organisms. Various reviews have described the range of assays that have been used for this purpose. Nevertheless, there continues to be confusion regarding acceptable methods to measure autophagy, especially in multicellular eukaryotes. A key point that needs to be emphasized is that there is a difference between measurements that monitor the numbers or volume of autophagic elements (e.g., autophagosomes or autolysosomes) at any stage of the autophagic process vs. those that measure flux through the autophagy pathway (i.e., the complete process); thus, a block in macroautophagy that results in autophagosome accumulation needs to be differentiated from stimuli that result in increased autophagic activity, defined as increased autophagy induction coupled with increased delivery to, and degradation within, lysosomes (in most higher eukaryotes and some protists such as Dictyostelium) or the vacuole (in plants and fungi). In other words, it is especially important that investigators new to the field understand that the appearance of more autophagosomes does not necessarily equate with more autophagy. In fact, in many cases, autophagosomes accumulate because of a block in trafficking to lysosomes without a concomitant change in autophagosome biogenesis, whereas an increase in autolysosomes may reflect a reduction in degradative activity. Here, we present a set of guidelines for the selection and interpretation of methods for use by investigators who aim to examine macroautophagy and related processes, as well as for reviewers who need to provide realistic and reasonable critiques of papers that are focused on these processes. These guidelines are not meant to be a formulaic set of rules, because the appropriate assays depend in part on the question being asked and the system being used. In addition, we emphasize that no individual assay is guaranteed to be the most appropriate one in every situation, and we strongly recommend the use of multiple assays to monitor autophagy. In these guidelines, we consider these various methods of assessing autophagy and what information can, or cannot, be obtained from them. Finally, by discussing the merits and limits of particular autophagy assays, we hope to encourage technical innovation in the field.

Recent research into various aspects of bacterial metabolism such as cell wall and antibiotic synthesis, degradation pathways, cellular stress, and amino acid biosynthesis has elucidated roles of aminoacyl‐transfer ribonucleic acid (aa‐tRNA) outside of translation. Although the two enzyme families responsible for cell wall modifications, aminoacyl‐phosphatidylglycerol synthases (aaPGSs) and Fem, were discovered some time ago, they have recently become of intense interest for their roles in the antimicrobial resistance of pathogenic microorganisms. The addition of positively charged amino acids to phosphatidylglycerol (PG) by aaPGSs neutralizes the lipid bilayer making the bacteria less susceptible to positively charged antimicrobial agents. Fem transferases utilize aa‐tRNA to form peptide bridges that link strands of peptidoglycan. These bridges vary among the bacterial species in which they are present and play a role in resistance to antibiotics that target the cell wall. Additionally, the formation of truncated peptides results in shorter peptide bridges and loss of branched linkages which makes bacteria more susceptible to antimicrobials. A greater understanding of the structure and substrate specificity of this diverse enzymatic family is necessary to aid current efforts in designing potential bactericidal agents. These two enzyme families are linked only by the substrate with which they modify the cell wall, aa‐tRNA; their structure, cell wall modification processes and the physiological changes they impart on the bacterium differ greatly. WIREs RNA2012, 3:247–264. doi: 10.1002/wrna.1108