The NADPH oxidase 2 subunit p47phox binds to the WAVE regulatory complex and p22phox in a mutually exclusive manner.

The actin cytoskeleton and reactive oxygen species (ROS) both play crucial roles in various cellular processes. Previous research indicated a direct interaction between two key components of these systems: the WAVE1 subunit of the WAVE regulatory complex (WRC), which promotes actin polymerization and the p47phox subunit of the NADPH oxidase 2 complex (NOX2), which produces ROS. Here, using carefully characterized recombinant proteins, we find that activated p47phox uses its dual Src homology 3 domains to bind to multiple regions within the WAVE1 and Abi2 subunits of the WRC, without altering WRC's activity in promoting Arp2/3-mediated actin polymerization. Notably, contrary to previous findings, p47phox uses the same binding pocket to interact with both the WRC and the p22phox subunit of NOX2, albeit in a mutually exclusive manner. This observation suggests that when activated, p47phox may separately participate in two distinct processes: assembling into NOX2 to promote ROS production and engaging with WRC to regulate the actin cytoskeleton.

The Toxoplasma secreted effector TgWIP modulates dendritic cell motility by activating host tyrosine phosphatases Shp1 and Shp2.

The obligate intracellular parasite Toxoplasma gondii causes life-threatening toxoplasmosis to immunocompromised individuals. The pathogenesis of Toxoplasma relies on its swift dissemination to the central nervous system through a 'Trojan Horse' mechanism using infected leukocytes as carriers. Previous work found TgWIP, a protein secreted from Toxoplasma, played a role in altering the actin cytoskeleton and promoting cell migration in infected dendritic cells (DCs). However, the mechanism behind these changes was unknown. Here, we report that TgWIP harbors two SH2-binding motifs that interact with tyrosine phosphatases Shp1 and Shp2, leading to phosphatase activation. DCs infected with Toxoplasma exhibited hypermigration, accompanying enhanced F-actin stress fibers and increased membrane protrusions such as filopodia and pseudopodia. By contrast, these phenotypes were abrogated in DCs infected with Toxoplasma expressing a mutant TgWIP lacking the SH2-binding motifs. We further demonstrated that the Rho-associated kinase (Rock) is involved in the induction of these phenotypes, in a TgWIP-Shp1/2 dependent manner. Collectively, the data uncover a molecular mechanism by which TgWIP modulates the migration dynamics of infected DCs in vitro.

On synergy between ultrahigh throughput screening and machine learning in biocatalyst engineering.

Protein design and directed evolution have separately contributed enormously to protein engineering. Without being mutually exclusive, the former relies on computation from first principles, while the latter is a combinatorial approach based on chance. Advances in ultrahigh throughput (uHT) screening, next generation sequencing and machine learning may create alternative routes to engineered proteins, where functional information linked to specific sequences is interpreted and extrapolated in silico. In particular, the miniaturisation of functional tests in water-in-oil emulsion droplets with picoliter volumes and their rapid generation and analysis (>1 kHz) allows screening of >107-membered libraries in a day. Subsequently, decoding the selected clones by short or long-read sequencing methods leads to large sequence-function datasets that may allow extrapolation from experimental directed evolution to further improved mutants beyond the observed hits. In this work, we explore experimental strategies for how to draw up 'fitness landscapes' in sequence space with uHT droplet microfluidics, review the current state of AI/ML in enzyme engineering and discuss how uHT datasets may be combined with AI/ML to make meaningful predictions and accelerate biocatalyst engineering.

Manipulating stereoselectivity of parahydrogen addition to acetylene to unravel interconversion of ethylene nuclear spin isomers.

Symmetric molecules exist as distinct nuclear spin isomers (NSIMs). A deeper understanding of their properties, including interconversion of different NSIMs, requires efficient techniques for NSIM enrichment. In this work, selective hydrogenation of acetylene with parahydrogen (p-H2) was used to achieve the enrichment of ethylene NSIMs and to study their equilibration processes. The effect of the stereoselectivity of H2 addition to acetylene on the imbalance of ethylene NSIMs was experimentally demonstrated by using three different heterogeneous catalysts (an immobilized Ir complex and two supported Pd catalysts). The interconversion of NSIMs with time during ethylene storage was studied using NMR spectroscopy by reacting ethylene with bromine water, which rendered the p-H2-derived protons in the produced 2-bromoethan(2H)ol (BrEtOD) magnetically inequivalent, thereby revealing the non-equilibrium nuclear spin order of ethylene. A thorough analysis of the shape and transformation of the 1H NMR spectra of hyperpolarized BrEtOD allowed us to reveal the initial distribution of produced ethylene NSIMs and their equilibration processes. Comparison of the results obtained with three different catalysts was key to properly attributing the derived characteristic time constants to different ethylene NSIM interconversion processes: ∼3-6 s for interconversion between NSIMs with the same inversion symmetry (i.e., within g or u manifolds) and ∼1700-2200 s between NSIMs with different inversion symmetries (i.e., between g and u manifolds).

Ligament augmentation and reconstruction system (LARS) synthetic grafts are safe and effective for medial collateral ligament and posterolateral corner reconstructions in elite athletes.

PURPOSE: This study documents the efficacy and safety of using a Ligament Augmentation and Reconstruction System (LARS) ligament graft to augment extra-articular knee ligament reconstructions in elite athletes by reporting return-to-play (RTP) rates and levels, career longevity and complications. METHODS: A consecutive series of all extra-articular knee ligament reconstructions augmented by LARS ligaments in elite athletes undertaken by three specialist sports knee surgeons between 2013 and 2020 were reviewed. Seventy-six elite athletes, aged over 16 years old, and more than 2 years postsurgery were included. RTP was defined as competing at professional level or national/international level in amateur sport. RESULTS: There were 64 medial collateral ligament (MCL) and 12 posterolateral corner (PLC) reconstructions. Fifty-two (68.4%) underwent concomitant autograft cruciate ligament(s) reconstruction. The mean age was 25.1 years (SD ± 4.5). Most were football (35, 46.1%) or rugby players (35, 46.1%). Sixty-seven athletes (88.2%) RTP with 65 (97.0%) of these playing at the same or higher Tegner level. Fifty-six (83.6%) of the athletes that RTP were still playing at 2 years postsurgery. Twenty (57.1%) of those who had RTP and were more than 5 years postsurgery were still playing at 5 years. Six (7.9%) players required further surgery relating to the LARS/metalwork. One case had soft tissue inflammation adjacent to the proximal end of the synthetic graft, but it is unknown if this was mechanical irritation or a biological reaction. One MCL reruptured 4 years after RTP. CONCLUSION: Utilising LARS to augment extra-articular knee ligament reconstructions allows 88.2% of athletes with a variety of knee ligament injuries to return to elite sport. The low morbidity rates coupled with 57% of athletes still playing 5 years postsurgery demonstrates that the LARS is safe and effective in these cases. Although there are reports of LARS ligaments being used in MCL and PLC reconstructions, there is very little evidence investigating if they are safe and effective. This study demonstrates that LARS synthetic grafts can be safely used for MCL and PLC reconstructions in elite athletes and they permit a high RTP with a low risk of complications. LEVEL OF EVIDENCE: Level IV.

The Influence of Heteroresistance on Minimum Inhibitory Concentration, Investigated Using Weak-Acid Stress in Food Spoilage Yeasts.

Populations of microbial cells may resist environmental stress by maintaining a high population-median resistance (IC50) or, potentially, a high variability in resistance between individual cells (heteroresistance); where heteroresistance would allow certain cells to resist high stress, provided the population was sufficiently large to include resistant cells. This study sets out to test the hypothesis that both IC50 and heteroresistance may contribute to conventional minimal inhibitory concentration (MIC) determinations, using the example of spoilage-yeast resistance to the preservative sorbic acid. Across a panel of 26 diverse yeast species, both heteroresistance and particularly IC50 were positively correlated with predicted MIC. A focused panel of 29 different isolates of a particular spoilage yeast was also examined (isolates previously recorded as Zygosaccharomyces bailii, but genome resequencing revealing that several were in fact hybrid species, Z. parabailii and Z. pseudobailii). Applying a novel high-throughput assay for heteroresistance, it was found that IC50 but not heteroresistance was positively correlated with predicted MIC when considered across all isolates of this panel, but the heteroresistance-MIC interaction differed for the individual Zygosaccharomyces subspecies. Z. pseudobailii exhibited higher heteroresistance than Z. parabailii whereas the reverse was true for IC50, suggesting possible alternative strategies for achieving high MIC between subspecies. This work highlights the limitations of conventional MIC measurements due to the effect of heteroresistance in certain organisms, as the measured resistance can vary markedly with population (inoculum) size. IMPORTANCE Food spoilage by fungi is a leading cause of food waste, with specialized food spoilage yeasts capable of growth at preservative concentrations above the legal limit, in part due to heteroresistance allowing small subpopulations of cells to exhibit extreme preservative resistance. Whereas heteroresistance has been characterized in numerous ecological contexts, measuring this phenotype systematically and assessing its importance are not encompassed by conventional assay methods. The development here of a high-throughput method for measuring heteroresistance, amenable to automation, addresses this issue and has enabled characterization of the contribution that heteroresistance may make to conventional MIC measurements. We used the example of sorbic acid heteroresistance in spoilage yeasts like Zygosaccharomyces spp., but the approach is relevant to other fungi and other inhibitors, including antifungals. The work shows how median resistance, heteroresistance, and inoculum size should all be considered when selecting appropriate inhibitor doses in real-world antimicrobial applications such as food preservation.

Changes in circulating forms of anti-Muüllerian hormone and androgens in women with and without PCOS: a systematic longitudinal study throughout pregnancy.

STUDY QUESTION: What are the changes in serum concentration of total and cleaved anti-Muüllerian hormone (AMH) molecular forms and of androgens before and throughout pregnancy in women with and without polycystic ovary syndrome (PCOS) in a longitudinal follow-up investigation? SUMMARY ANSWER: Serum levels of total and cleaved AMH are higher from preconception to the third trimester of pregnancy in women with PCOS as compared to controls, whereas testosterone and androstenedione levels are higher in women with PCOS than in control women before pregnancy and during the second and third trimester of pregnancy. WHAT IS KNOWN ALREADY: Cross-sectional or partial longitudinal studies have shown higher AMH and androgen levels in pregnant women with PCOS as compared with non-PCOS women. To date, no complete longitudinal dynamic monitoring of the circulating forms of AMH and androgens from pre-conception to the third trimester of pregnancy have compared women with and without PCOS. STUDY DESIGN, SIZE, DURATION: This systematic prospective quarterly longitudinal monocentric study was a comparative follow-up of 30 women with PCOS and 29 controls before and during pregnancy from April 2019 to July 2022. PARTICIPANTS/MATERIALS, SETTING, METHODS: Women aged 18-43 years with a pre-conception measurement of AMH were included during the first trimester of a singleton pregnancy. The PCOS group was defined according to the Rotterdam diagnostic criteria. The control group patients included in the study had normal ovarian reserves. Circulating total and cleaved AMH, and serum estradiol, LH, and androgen levels were measured during the first, second, and third trimester of pregnancy in all study participants. MAIN RESULTS AND THE ROLE OF CHANCE: Before pregnancy, patients with PCOS had higher levels of AMH than controls. The total and cleaved AMH forms were significantly higher in women with PCOS than controls from pre-conception to the third trimester of pregnancy (all P < 0.001). Androgens (total testosterone and androstenedione) were higher in women with PCOS than controls from mid-pregnancy onwards. LIMITATIONS, REASONS FOR CAUTION: Our control population was a population of infertile women with no ovarian problems but most of them had undergone ART treatments to achieve pregnancy. WIDER IMPLICATIONS OF THE FINDINGS: These results strengthen the hypothesis that gestational hyperandrogenism as well as exposure to elevated AMH levels in utero could be driving forces predisposing female progeny to develop PCOS. STUDY FUNDING/COMPETING INTEREST(S): Funding was provided by INSERM, France (grant number U1172) and the European Research Council (ERC) under the European Union's Horizon 2020 research and innovation program, ERC-2016-CoG to P.G. grant agreement n° 725149/REPRODAMH. The authors have nothing to declare. TRIAL REGISTRATION NUMBER: NCT03483792.

Clonal dynamics in pediatric B-cell precursor acute lymphoblastic leukemia with very early relapse.

INTRODUCTION: One-quarter of the relapses in children with B-cell precursor acute lymphoblastic leukemia (BCP-ALL) occur very early (within 18 months, before completion of treatment), and prognosis in these patients is worse compared to cases that relapse after treatment has ended. METHODS: In this study, we performed a genomic analysis of diagnosis-relapse pairs of 12 children who relapsed very early, followed by a deep-sequencing validation of all identified mutations. In addition, we included one case with a good initial treatment response and on-treatment relapse at the end of upfront therapy. RESULTS: We observed a dynamic clonal evolution in all cases, with relapse almost exclusively originating from a subclone at diagnosis. We identified several driver mutations that may have influenced the outgrowth of a minor clone at diagnosis to become the major clone at relapse. For example, a minimal residual disease (MRD)-based standard-risk patient with ETV6-RUNX1-positive leukemia developed a relapse from a TP53-mutated subclone after loss of the wildtype allele. Furthermore, two patients with TCF3-PBX1-positive leukemia that developed a very early relapse carried E1099K WHSC1 mutations at diagnosis, a hotspot mutation that was recurrently encountered in other very early TCF3-PBX1-positive leukemia relapses as well. In addition to alterations in known relapse drivers, we found two cases with truncating mutations in the cohesin gene RAD21. CONCLUSION: Comprehensive genomic characterization of diagnosis-relapse pairs shows that very early relapses in BCP-ALL frequently arise from minor subclones at diagnosis. A detailed understanding of the therapeutic pressure driving these events may aid the development of improved therapies.

Microbial Metal Resistance within Structured Environments Is Inversely Related to Environmental Pore Size.

The physical environments in which microorganisms naturally reside rarely have homogeneous structure, and changes in their porous architecture may have effects on microbial activities that are not typically captured in conventional laboratory studies. In this study, to investigate the influence of environmental structure on microbial responses to stress, we constructed structured environments with different pore properties (determined by X-ray computed tomography). First, using glass beads in different arrangements and inoculated with the soil yeast Saitozyma podzolica, increases in the average equivalent spherical diameters (ESD) of a structure's porous architecture led to decreased survival of the yeast under a toxic metal challenge with lead nitrate. This relationship was reproduced when yeasts were introduced into additively manufactured lattice structures, comprising regular arrays with ESDs comparable to those of the bead structures. The pore ESD dependency of metal resistance was not attributable to differences in cell density in microenvironments delimited by different pore sizes, supporting the inference that pore size specifically was the important parameter in determining survival of stress. These findings highlight the importance of the physical architecture of an organism's immediate environment for its response to environmental perturbation, while offering new tools for investigating these interactions in the laboratory. IMPORTANCE Interactions between cells and their structured environments are poorly understood but have significant implications for organismal success in both natural and nonnatural settings. This work used a multidisciplinary approach to develop laboratory models with which the influence of a key parameter of environmental structure-pore size-on cell activities can be dissected. Using these new methods in tandem with additive manufacturing, we demonstrated that resistance of yeast soil isolates to stress (from a common metal pollutant) is inversely related to pore size of their environment. This has important ramifications for understanding how microorganisms respond to stress in different environments. The findings also establish new pathways for resolving the effects of physical environment on microbial activity, enabling important understanding that is not readily attainable with traditional bulk sampling and analysis approaches.

Multiclonal complexity of pediatric acute lymphoblastic leukemia and the prognostic relevance of subclonal mutations

Genomic studies of pediatric acute lymphoblastic leukemia (ALL) have shown remarkable heterogeneity in initial diagnosis, with multiple (sub)clones harboring lesions in relapse-associated genes. However, the clinical relevance of these subclonal alterations remains unclear. We assessed the clinical relevance and prognostic value of subclonal alterations in the relapse-associated genes IKZF1, CREBBP, KRAS, NRAS, PTPN11, TP53, NT5C2, and WHSC1 in 503 ALL cases. Using Molecular Inversion Probe sequencing and breakpoint-spanning PCR we reliably detected alterations below 1% allele frequency. We identified 660 genomic alterations in 285 diagnosis samples of which 495 (75%) were subclonal. RAS pathway mutations were common, particularly in minor subclones, and comparisons between RAS hotspot mutations revealed differences in their capacity to drive clonal expansion in ALL. We did not find an association of subclonal alterations with unfavorable outcome. Particularly for IKZF1, an established prognostic marker in ALL, all clonal but none of the subclonal alterations were preserved at relapse. We conclude that, for the genes tested, there is no basis to consider subclonal alterations detected at diagnosis for risk group stratification of ALL treatment.

Potentiated inhibition of Trichoderma virens and other environmental fungi by new biocide combinations.

Fungi cause diverse, serious socio-economic problems, including biodeterioration of valuable products and materials that spawns a biocides industry worth ~$11 billion globally. To help combat environmental fungi that commonly colonise material products, this study tested the hypothesis that combination of an approved fungicide with diverse agents approved by the FDA (Food and Drug Administration) could reveal potent combinatorial activities with promise for fungicidal applications. The strategy to use approved compounds lowers potential development risks for any effective combinations. A high-throughput assay of 1280 FDA-approved compounds was conducted to find those that potentiate the effect of iodopropynyl-butyl-carbamate (IPBC) on the growth of Trichoderma virens; IPBC is one of the two most widely used Biocidal Products Regulations-approved fungicides. From this library, 34 compounds in combination with IPBC strongly inhibited fungal growth. Low-cost compounds that gave the most effective growth inhibition were tested against other environmental fungi that are standard biomarkers for resistance of synthetic materials to fungal colonisation. Trifluoperazine (TFZ) in combination with IPBC enhanced growth inhibition of three of the five test fungi. The antifungal hexetidine (HEX) potentiated IPBC action against two of the test organisms. Testable hypotheses on the mechanisms of these combinatorial actions are discussed. Neither IPBC + TFZ nor IPBC + HEX exhibited a combinatorial effect against mammalian cells. These combinations retained strong fungal growth inhibition properties after incorporation to a polymer matrix (alginate) with potential for fungicide delivery. The study reveals the potential of such approved compounds for novel combinatorial applications in the control of fungal environmental opportunists. KEY POINTS: • Search with an approved fungicide to find new fungicidal synergies in drug libraries. • New combinations inhibit growth of key environmental fungi on different matrices. • The approach enables a more rapid response to demand for new biocides.

Should We Abandon Systematic Pelvic and Paraaortic Lymphadenectomy in Low-Grade Serous Ovarian Cancer?

BACKGROUND: Low-grade serous ovarian carcinoma (LGSOC) is a rare disease that accounts for 5% of all ovarian cancers and requires surgical complete debulking. To date, the prognostic value of pelvic and paraaortic lymphadenectomy remains unclear in this population. PATIENTS AND METHODS: This retrospective cohort of patients with a diagnosis of LGSOC was registered in the Tumeurs Malignes Rares Gynécologiques national network, between January 2000 and July 2017, at 25 centers. All LGSOC were confirmed after pathological review and operated by primary debulking surgery (PDS) or interval debulking surgery after neoadjuvant chemotherapy (NACT-IDS). Primary endpoints were overall survival (OS) and progression-free survival (PFS). RESULTS: A total of 126 patients were included, 86.1% were stage III/IV, and 74.6% underwent lymph node dissection (LND). According to the Completeness of Cancer Resection (CCR) score, 83.7% had complete resection. Median OS was 130 months, and median PFS was 41 months. Pelvic and paraaortic LND had no significant impact on OS (p = 0.78) or DFS (p = 0.93), and this was confirmed in subgroups (advanced stages FIGO III/IV, CCR score 0/1 or 2/3, and timing of surgery PDS or NACT-IDS). Histological positive paraaortic lymph nodes had a significant negative impact on PFS in the whole population (HR 2.21, 1.18-4.39, p = 0.02) and in the CC0/CC1 population (HR, 2.28, 1.13-4.59, p = 0.02). CONCLUSIONS: Systematic pelvic and paraaortic LND in patients with LGSOC improved neither overall nor PFS. A prospective trial would be necessary to validate these results but would be difficult to conduct due to the rarity of this disease.

The Ca(v)1.4 calcium channel is a critical regulator of T cell receptor signaling and naive T cell homeostasis.

The transport of calcium ions (Ca(2+)) to the cytosol is essential for immunoreceptor signaling, regulating lymphocyte differentiation, activation, and effector function. Increases in cytosolic-free Ca(2+) concentrations are thought to be mediated through two interconnected and complementary mechanisms: the release of endoplasmic reticulum Ca(2+) "stores" and "store-operated" Ca(2+) entry via plasma membrane channels. However, the identity of molecular components conducting Ca(2+) currents within developing and mature T cells is unclear. Here, we have demonstrated that the L-type "voltage-dependent" Ca(2+) channel Ca(V)1.4 plays a cell-intrinsic role in the function, development, and survival of naive T cells. Plasma membrane Ca(V)1.4 was found to be essential for modulation of intracellular Ca(2+) stores and T cell receptor (TCR)-induced rises in cytosolic-free Ca(2+), impacting activation of Ras-extracellular signal-regulated kinase (ERK) and nuclear factor of activated T cells (NFAT) pathways. Collectively, these studies revealed that Ca(V)1.4 functions in controlling naive T cell homeostasis and antigen-driven T cell immune responses.

Presence of Genetic Variants Among Young Men With Severe COVID-19.

Importance: Severe coronavirus disease 2019 (COVID-19) can occur in younger, predominantly male, patients without preexisting medical conditions. Some individuals may have primary immunodeficiencies that predispose to severe infections caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Objective: To explore the presence of genetic variants associated with primary immunodeficiencies among young patients with COVID-19. Design, Setting, and Participants: Case series of pairs of brothers without medical history meeting the selection criteria of young (age <35 years) brother pairs admitted to the intensive care unit (ICU) due to severe COVID-19. Four men from 2 unrelated families were admitted to the ICUs of 4 hospitals in the Netherlands between March 23 and April 12, 2020. The final date of follow-up was May 16, 2020. Available family members were included for genetic variant segregation analysis and as controls for functional experiments. Exposure: Severe COVID-19. Main Outcome and Measures: Results of rapid clinical whole-exome sequencing, performed to identify a potential monogenic cause. Subsequently, basic genetic and immunological tests were performed in primary immune cells isolated from the patients and family members to characterize any immune defects. Results: The 4 male patients had a mean age of 26 years (range, 21-32), with no history of major chronic disease. They were previously well before developing respiratory insufficiency due to severe COVID-19, requiring mechanical ventilation in the ICU. The mean duration of ventilatory support was 10 days (range, 9-11); the mean duration of ICU stay was 13 days (range, 10-16). One patient died. Rapid clinical whole-exome sequencing of the patients and segregation in available family members identified loss-of-function variants of the X-chromosomal TLR7. In members of family 1, a maternally inherited 4-nucleotide deletion was identified (c.2129_2132del; p.[Gln710Argfs*18]); the affected members of family 2 carried a missense variant (c.2383G>T; p.[Val795Phe]). In primary peripheral blood mononuclear cells from the patients, downstream type I interferon (IFN) signaling was transcriptionally downregulated, as measured by significantly decreased mRNA expression of IRF7, IFNB1, and ISG15 on stimulation with the TLR7 agonist imiquimod as compared with family members and controls. The production of IFN-γ, a type II IFN, was decreased in patients in response to stimulation with imiquimod. Conclusions and Relevance: In this case series of 4 young male patients with severe COVID-19, rare putative loss-of-function variants of X-chromosomal TLR7 were identified that were associated with impaired type I and II IFN responses. These preliminary findings provide insights into the pathogenesis of COVID-19.

Stereoselectivity of Electron and Energy Transfer in the Quenching of (S/R)-Ketoprofen-(S)-Tryptophan Dyad Excited State.

Photoinduced elementary processes in chiral linked systems, consisting of drugs and tryptophan (Trp) residues, attract considerable attention due to several aspects. First of all, these are models that allow one to trace the full and partial charge transfer underlying the binding of drugs to enzymes and receptors. On the other hand, Trp fluorescence is widely used to establish the structure and conformational mobility of proteins due to its high sensitivity to the microenvironment. Therefore, the study of mechanisms of Trp fluorescence quenching in various systems has both fundamental and practical interest. An analysis of the photo-chemically induced dynamic nuclear polarization (CIDNP) and Trp fluorescence quenching in (R/S)-ketoprofen-(S)-tryptophan ((S/R)-KP-(S)-Trp) dyad carried out in this work allowed us to trace the intramolecular reversible electron transfer (ET) and obtain evidence in favor of the resonance energy transfer (RET). The fraction of dyad's singlet excited state, quenched via ET, was shown to be 7.5 times greater for the (S,S)-diastereomer than for the (R,S) analog. At the same time, the ratio of the fluorescence quantum yields shows that quenching effectiveness of (S,S)-diastereomer to be 5.4 times lower than for the (R,S) analog. It means that the main mechanism of Trp fluorescence quenching in (S/R)-KP-(S)-Trp dyad is RET.

TRIM28 haploinsufficiency predisposes to Wilms tumor.

Two percent of patients with Wilms tumors have a positive family history. In many of these cases the genetic cause remains unresolved. By applying germline exome sequencing in two families with two affected individuals with Wilms tumors, we identified truncating mutations in TRIM28. Subsequent mutational screening of germline and tumor DNA of 269 children affected by Wilms tumor was performed, and revealed seven additional individuals with germline truncating mutations, and one individual with a somatic truncating mutation in TRIM28. TRIM28 encodes a complex scaffold protein involved in many different processes, including gene silencing, DNA repair and maintenance of genomic integrity. Expression studies on mRNA and protein level showed reduction of TRIM28, confirming a loss-of-function effect of the mutations identified. The tumors showed an epithelial-type histology that stained negative for TRIM28 by immunohistochemistry. The tumors were bilateral in six patients, and 10/11 tumors are accompanied by perilobar nephrogenic rests. Exome sequencing on eight tumor DNA samples from six individuals showed loss-of-heterozygosity (LOH) of the TRIM28-locus by mitotic recombination in seven tumors, suggesting that TRIM28 functions as a tumor suppressor gene in Wilms tumor development. Additionally, the tumors showed very few mutations in known Wilms tumor driver genes, suggesting that loss of TRIM28 is the main driver of tumorigenesis. In conclusion, we identified heterozygous germline truncating mutations in TRIM28 in 11 children with mainly epithelial-type Wilms tumors, which become homozygous in tumor tissue. These data establish TRIM28 as a novel Wilms tumor predisposition gene, acting as a tumor suppressor gene by LOH.

Osteoporosis prevention: where are the barriers to improvement in a French general population? A qualitative study.

We conducted a qualitative study with French men and women in order to provide insight into individuals' experiences, behaviors, and perceptions about osteoporosis (OP) and OP care. The data showed that both sexes, but especially men, were unfamiliar with OP, did not always feel concerned, and mistrusted pharmacological treatments. INTRODUCTION: To engage actively in osteoporosis (OP) prevention, people need to have basic knowledge about the disease. The aim of this qualitative study was to explore knowledge and representations of OP care and prevention among both men and women. METHODS: Focus groups were conducted in the Rhône-Alpes Region, France, with women aged 50-85 years and men aged 60-85 years, with or without a history of fragility fracture and/or an OP diagnosis (respectively referred to as "aware" or "unaware"). A total of 45 women (23 "aware" and 22 "unaware" in 5 and 4 focus groups, respectively) and 53 men (19 "aware" and 34 "unaware" in 3 and 4 focus groups, respectively) were included. A thematic analysis of transcripts was performed to explore knowledge and representations about OP, risk factors, prevention, and treatment. RESULTS: The data showed that both sexes, but especially men, had limited knowledge of OP and considered it as a natural aging process not related to fragility fractures. They generally did not feel concerned by OP and no important difference was observed between "aware" and "unaware" patients. Women expressed their fear of the disease, associated with aging and the end of life, while men considered it to be a women's disease only. Both sexes were aware of OP risk factors, but were suspicious towards treatments because of the associated side effects. CONCLUSION: Understanding people's representation of OP might help to provide patients with relevant information in order to optimize their preventive behavior and decrease the burden of the disease.

What goes up must come down: biomechanical impact analysis of falling locusts.

Many insects are able to precisely control their jumping movements. Once in the air, the properties of the actual landing site, however, are almost impossible to predict. Falling insects thus have to cope with the situation at impact. In particular, for insects jumping to escape predators, a controlled landing movement appears to be a major evolutionary advantage. A quick recovery into an upright and stable body posture minimizes the time to prepare for the next escape jump. In this study, we used high-speed recordings to investigate the falling and in particular the impact behavior of Schistocerca gregaria locusts, a common model organism for studies on the biomechanics of jumping. Detailed impact analyses of free-falling locusts show that most insects typically crashed onto the substrate. Although free-falling locusts tended to spread their legs, they mostly fell onto the head and thorax first. The presence of wings did not significantly reduce impact speed; however, it did affect the orientation of the body at impact and significantly reduced the time to recover. Our results also show that alive warm locusts fell significantly faster than inactive or dead locusts. This indicates a possible tradeoff between active control versus reduced speed. Interestingly, alive insects also tended to perform a characteristic bending movement of the body at impact. This biomechanical adaptation might reduce the rebound and shorten the time to recover. The adhesive pads also play an important role in reducing the time to recover by allowing the insect to anchor itself to the substrate.

Microbes associated with fresh produce: Sources, types and methods to reduce spoilage and contamination.

Global food security remains one of the most important challenges that needs to be addressed to ensure the increasing demand for food of the fast growing human population is satisfied. Fruits and vegetables comprise an essential component of a healthy balanced diet as they are the major source of both macro- and micronutrients. They are particularly important for communities in developing countries whose nutrition often relies solely on a plant-based diet. Recent advances in agriculture and food processing technologies have facilitated production of fresh, nutritious and safe food for consumers. However, despite the development of sophisticated chemical and physical methods of food and equipment disinfection, fresh-cut produce and fruit juice industry still faces significant economic losses due to microbial spoilage. Furthermore, fresh produce remains an important source of pathogens that have been causing outbreaks of human illness worldwide. This chapter characterizes common spoilage and human pathogenic microorganisms associated with fresh-cut produce and fruit juice products, and discusses the methods and technology that have been developed and utilized over the years to combat them. Substantial attention is given to highlight advantages and disadvantages of using these methods to reduce microbial spoilage and their efficacy to eliminate human pathogenic microbes associated with consumption of fresh-cut produce and fruit juice products.

Effects of 5-ALA dose on resection of glioblastoma.

PURPOSE: Fluorescence-guided surgery (FGS) with the use of 5-aminolevulinic acid (5-ALA) leads to more extensive resection of high-grade glioma (HGG) and longer overall survival (OS) of patients compared to conventional resection. The purpose of this study is to investigate the effect of 5-ALA dosages on residual tumor volume (RTV) and OS in patients with glioblastoma. METHODS: A retrospective cohort study for patients who participated in a phase I and II dose-escalation clinical trial on 5-ALA for resection of HGG. A total of 25 patients were found to have newly diagnosed glioblastoma on histology and enrolled in our study. Patients receiving low doses of 5-ALA (10-30 mg/kg) (n = 6) were compared to those receiving high doses (40-50 mg/kg) (n = 19). Pre- and post-operative contrast enhanced T1W MRI were evaluated with volumetric analysis. RESULTS: Median RTV was 0.69 cm3 and 0.00 cm3 in the low and high dose groups respectively (p = 0.975). A gross total resection (GTR) was more likely in the high dose group, though this was not statistically significant. No significant difference was found in median OS between the high and low dose groups (p = 0.6787). CONCLUSIONS: High doses of 5-ALA FGS are associated with less RTV and greater probability of GTR. 5-ALA dose was not associated with OS. Further studies with a larger patient cohort are warranted.