In situ crosslinkable multi-functional and cell-responsive alginate 3D matrix via thiol-maleimide click chemistry.

In vivo, cells interact with the extracellular matrix (ECM), which provides a multitude of biophysical and biochemical signals that modulate cellular behavior. Inspired by this, we explored a new methodology to develop a more physiomimetic polysaccharide-based matrix for 3D cell culture. Maleimide-modified alginate (AlgM) derivatives were successfully synthesized using DMTMM to activate carboxylic groups. Thiol-terminated cell-adhesion peptides were tethered to the hydrogel network to promote integrin binding. Rapid and efficient in situ hydrogel formation was promoted by thiol-Michael addition "click" chemistry via maleimide reaction with thiol-flanked protease-sensitive peptides. Alginate derivatives were further ionically crosslinked by divalent ions present in the medium, which led to greater stability and allowed longer cell culture periods. By tailoring alginate's biofunctionality we improved cell-cell and cell-matrix interactions, providing an ECM-like 3D microenvironment. We were able to systematically and independently vary biochemical and biophysical parameters to elicit specific cell responses, creating custom-made 3D matrices. DMTMM-mediated maleimide incorporation is a promising approach to synthesizing AlgM derivatives that can be leveraged to produce ECM-like matrices for a broad range of applications, from in vitro tissue modeling to tissue regeneration.

Sex-Specific Patterns of Cerebral Atrophy and Enlarged Perivascular Spaces in Patients with Cerebral Amyloid Angiopathy and Dementia.

BACKGROUND AND PURPOSE: Cerebral amyloid angiopathy is characterized by amyloid ß deposition in leptomeningeal and superficial cortical vessels. Cognitive impairment is common and may occur independent of concomitant Alzheimer disease neuropathology. It is still unknown which neuroimaging findings are associated with dementia in cerebral amyloid angiopathy and whether they are modulated by sex. This study compared MR imaging markers in patients with cerebral amyloid angiopathy with dementia or mild cognitive impairment or who are cognitively unimpaired and explored sex-specific differences. MATERIALS AND METHODS: We studied 58 patients with cerebral amyloid angiopathy selected from the cerebrovascular and memory outpatient clinics. Clinical characteristics were collected from clinical records. Cerebral amyloid angiopathy was diagnosed on MR imaging on the basis of the Boston criteria. Visual rating scores for atrophy and other imaging features were independently assessed by 2 senior neuroradiologists. RESULTS: Medial temporal lobe atrophy was higher for those with cerebral amyloid angiopathy with dementia versus those cognitively unimpaired (P = .015), but not for those with mild cognitive impairment. This effect was mainly driven by higher atrophy in men with dementia, compared with women with and without dementia (P = .034, P = .012; respectively) and with men without dementia (P = .012). Enlarged perivascular spaces in the centrum semiovale were more frequent in women with dementia versus men with and without dementia (P = .021, P = .011; respectively) and women without dementia (P = .011). CONCLUSIONS: Medial temporal lobe atrophy was more prominent in men with dementia, whereas women showed a higher number of enlarged perivascular spaces in the centrum semiovale. Overall, this finding suggests differential pathophysiologic mechanisms with sex-specific neuroimaging patterns in cerebral amyloid angiopathy.

Refinements in embryo manipulation applied to CRISPR technology in livestock.

The implementation of CRISPR technology in large animals requires further improvements in embryo manipulation and transfer to be applied with commercial purposes. In this study we report (a) developmental competence of CRISPR/Cas microinjected zygotes subjected to in vitro culture in large scale programs in sheep; (b) pregnancy outcomes after early-stage (2-8-cell) embryo transfer into the oviduct or the uterine horn; and (c) embryo survival and birth rate after vitrification/warming of CRISPR/Cas microinjected zygotes. Experiment 1 consisted of a retrospective analysis to evaluate embryo developmental rate of in vitro produced zygotes subjected to CRISPR/Cas microinjection (n = 7,819) compared with a subset of non-microinjected zygotes (n = 701). Development rates to blastocyst on Day 6 were 20.0% for microinjected zygotes and 44.9% for non-injected zygotes (P < 0.05). In Experiment 2, CRISPR/Cas microinjected zygotes were transferred on Day 2 after in vitro fertilization (2-8 cell embryos) into the oviductal ampulla (n = 262) or into the uterine horn (n = 276) in synchronized recipient ewes at prefixed time (i.e., approximately two days after ovulation). Pregnant/transferred recipients (24.0% vs. 25.0%), embryo survival/transferred embryos (6.9% vs. 6.2%), and born lambs/pregnant embryos (72.2% vs. 100.0%) did not differ significantly in the two groups. In Experiment 3, CRISPR/Cas microinjected zygotes were maintained under in vitro culture until blastocyst stage (Day 6), and subjected to vitrification/warming via the Cryotop method (n = 474), while a subset of embryos were left fresh as control group (n = 75). Embryos were transferred into the uterine horn of recipient females at prefixed time 8.5 days after the estrous synchronization treatment (i.e., approximately six days after ovulation). Pregnancy rate (30.8% vs. 48.0%), embryo survival rate (14.8% vs. 21.3%), and birth rate (85.7% vs. 75.0%) were not different (PNS) between vitrified and fresh embryos, respectively. In conclusion, the current study in sheep embryos reports (a) suitable developmental rate after CRISPR/Cas microinjection (i.e., 20%), even though it was lower than non-microinjected zygotes; (b) similar outcomes when Day 2-embryos were placed into the uterine horn instead of the oviduct, avoiding both time-consuming and invasive oviduct manipulation, and extended in vitro culture during one week; (c) promising pregnancy and birth rates obtained with vitrification of CRISPR/Cas microinjected embryos. This knowledge on in vitro embryo development, timing of embryo transfer, and cryopreservation of CRISPR/Cas microinjected zygotes have practical implications for the implementation of genome editing technology in large animals.

Starch-based films doped with porphyrinoid photosensitizers for active skin wound healing.

Starch is a biodegradable and biocompatible carbohydrate that, when combined with bioactive molecules, can be processed as biomimetic platforms with enhanced performance, allowing its use as active wound dressing materials. Porphyrinoid photosensitizers can tune the physicochemical/functional profile of biomacromolecules, allowing their use in anti-infective strategies. In this work, the feasibility of using the cationic 5,10,15,20-tetrakis(1-methylpyridinium-4-yl)porphyrin tetraiodide (TMPyP) to enhance the physicochemical, mechanical, antimicrobial performance, and wound healing ability of casted starch-based films was studied. TMPyP conferred a reddish coloration to the films, maintaining their pristine transparency. It increased by 87 % the films hydrophobicity and, depending on the TMPyP used, conferred mobility to the starch polymeric chains. Starch/TMPyP-based films effectively photoinactivated Escherichia coli (>99.99 %) and favored the wound healing process, even in the absence of light. Therefore, the incorporation of TMPyP into starch-based formulations revealed to be a promising strategy to tune the films compaction degree while giving rise to water tolerant and photosensitive biomaterials that can act as multitarget antimicrobial medical dressings and glycocarriers of active compounds relevant for effective skin wound healing.

Modeling the effectiveness of One Health interventions against the zoonotic hookworm Ancylostoma ceylanicum.

Hookworm disease is a major global public health concern, annually affecting 500-700 million of the world's poorest people. The World Health Organization is targeting the elimination of hookworm as a public health problem by 2030 using a strategy of mass drug administration (MDA) to at-risk human populations. However, in Southeast Asia and the Pacific the zoonotic hookworm species, Ancylostoma ceylanicum, is endemic in dogs and commonly infects people. This presents a potential impediment to the effectiveness of MDA that targets only humans. Here, we develop a novel multi-host (dog and human) transmission model of A. ceylanicum and compare the effectiveness of human-only and "One Health" (human plus dog) MDA strategies under a range of eco-epidemiological assumptions. We show that One Health interventions-targeting both dogs and humans-could suppress prevalence in humans to ≤ 1% by the end of 2030, even with only modest coverage (25-50%) of the animal reservoir. With increasing coverage, One Health interventions may even interrupt transmission. We discuss key unresolved questions on the eco-epidemiology of A. ceylanicum, the challenges of delivering MDA to animal reservoirs, and the growing importance of One Health interventions to human public health.

The impact of FSH stimulation and age on the ovarian and uterine traits and histomorphometry of prepubertal gilts.

This study investigated the effect of age and follicle stimulating hormone (FSH) treatment on the estradiol (E2) plasma concentration, ovarian follicle development, endometrial histomorphometry, and ultrasonographic parameters of the ovaries and uterus in prepubertal gilts. Thirty-five prepubertal gilts were grouped according to age (140 or 160 d), and within each age, gilts were allotted to receive 100 mg of FSH (treated; G140 + FSH [n = 10] and G160 + FSH [n = 7]) or saline solution (control; G140 + control [n = 10] and G160 + control [n = 8]). The total dose of FSH was divided into 6 similar doses administered every 8 h (days 0-2). Before and after FSH treatment, blood sample was collected, and transabdominal scanning of the ovaries and uterus was performed. Twenty-four hours after the last FSH injection, the gilts were slaughtered and their ovaries and uterus were processed for histological and histomorphometric analysis. The histomorphometric parameters of the uterus differed (P < 0.05) between prepubertal gilts at 160 d and 140 d of age. Moreover, changes (P < 0.05) in uterine and ovarian ultrasound images occurred between 140 and 160 d of age. Age and FSH treatment did not affect (P > 0.05) E2 plasma concentrations. Follicle stimulating hormone treatment did not affect (P > 0.05) the early stage of folliculogenesis in the prepubertal gilts; however, the number of early atretic follicles decreased (P < 0.05) after the FSH treatment. Follicle stimulating hormone administration increased (P < 0.05) the number of medium follicles and decreased (P < 0.05) the number of small follicles in 140 and 160 d old gilts. In the endometrium, luminal/glandular epithelium height and glandular diameter increased (P < 0.05) after FSH treatment. Thus, injections of 100 mg of FSH stimulate the endometrium epithelium and induce follicular growth to a medium follicle size without affecting the preantral stages in prepubertal gilts; also, the uterine macroscopic morphometry does not change from 140 to 160 d of age.

Safety assessment of new nanodiamonds@corrole hybrids addressed by the response of RAW-264.7 macrophages.

Safety assessment of carbon nanomaterials is of paramount importance since they are on the frontline for applications in sensing, bioimaging and drug delivery. The biocompatibility and safety of functionalized nanodiamonds (NDs) are here addressed through the study of the pro-inflammatory response of RAW-264.7 macrophages exposed to new nanodiamonds@corrole hybrids. The corrole unit selected is as a prototype for a hydrophobic organic molecule that can function as a NIR fluorophore reporter, an optical sensor, a photodynamic therapy agent or a photocatalyst. The new functional nanohybrids containing detonated nanodiamonds (NDs) were obtained through esterification using carboxylated NDs and glycol corroles. The success of the covalent functionalization via carbodiimide activation was confirmed through X-ray photoelectron spectroscopy (XPS), Raman and Fourier transform infrared (FTIR) spectroscopy. The UV-vis absorption and emission spectra of the hybrids are additive with respect to the corrole features. The cellular uptake, localization, cell viability and effects on immune cell activation of the new hybrids and of the precursors were carefully investigated using RAW-264.7 macrophages. Overall results showed that the ND@corrole hybrids had no pro-inflammatory effects on the RAW-264.7 macrophage cell line, making them an ideal candidate for a wide range of biomedical applications.

Impacts of the use of biological pest control on the technical efficiency of the Brazilian agricultural sector.

This paper aimed to assess the impact of the intensive use of biological control on the technical efficiency of the Brazilian agricultural sector. The study also considered the influence of factors such as technical assistance, rural financing, and membership in cooperatives or class entities on efficiency measures. It was estimated stochastic production frontiers for regions intensively using and not using biological control, considering potential selection bias. Results demonstrate that areas with intensive use of biological pest management have a 0.863 technical efficiency score, while this score is 0.823 for nonintensive areas. This means that the intensive regions would be closer to their efficiency frontiers. Additionally, technical assistance and membership in cooperative or class entities increase efficiency by 6 and 2.5%, respectively. It can be concluded that the intensive adoption of biological control can raise the productive performance of the Brazilian agricultural sector. Therefore, it must be highlighted the importance of formulating joint policies (e.g., credit+rural extension) for the adoption of biological control to be a feasible option to promote the sustainable development of the Brazilian agricultural sector.

End-to-end deep learning of geometric shaping for unamplified coherent systems.

With the increasing data rate requirements on short-reach links, the recent standardization of unamplified coherent optical systems is paving the way for a cost and power-effective solution, targeting a massive deployment in the near future. However, unamplified systems are introducing new challenges. Particularly, the performance is highly dependent on the peak-to-average power ratio (PAPR) of the transmitted signal, which puts at question the use of the typical constellation formats. In this work, we use an end-to-end deep learning framework to optimize the geometry of different constellation sizes, ranging from 8- to 128-ary constellations. In general, it is shown that the performance of these systems is maximized with constellations whose outer symbols are disposed in a square shape, owing to the minimization of the real-valued PAPR. Following this premise, we experimentally demonstrate that odd-bit constellations can be significantly optimized for unamplified coherent links, achieving power budget gains in the range of 0.5-3 dB through the geometric optimization of 8-, 32- and 128-ary constellations.

Potential drivers for schistosomiasis persistence: Population genetic analyses from a cluster-randomized urogenital schistosomiasis elimination trial across the Zanzibar islands.

The World Health Organization's revised NTD Roadmap and the newly launched Guidelines target elimination of schistosomiasis as a public health problem in all endemic areas by 2030. Key to meeting this goal is elucidating how selective pressures imposed by interventions shape parasite populations. Our aim was to identify any differential impact of a unique cluster-randomized tri-armed elimination intervention (biannual mass drug administration (MDA) applied alone or in association with either mollusciciding (snail control) or behavioural change interventions) across two Zanzibarian islands (Pemba and Unguja) on the population genetic composition of Schistosoma haematobium over space and time. Fifteen microsatellite loci were used to analyse individual miracidia collected from infected individuals across islands and intervention arms at the start (2012 baseline: 1,522 miracidia from 176 children; 303 from 43 adults; age-range 6-75, mean 12.7 years) and at year 5 (2016: 1,486 miracidia from 146 children; 214 from 25 adults; age-range 9-46, mean 12.4 years). Measures of genetic diversity included allelic richness (Ar), Expected (He) and Observed heterozygosity (Ho), inbreeding coefficient (FST), parentage analysis, estimated worm burden, worm fecundity, and genetic sub-structuring. There was little evidence of differential selective pressures on population genetic diversity, inbreeding or estimated worm burdens by treatment arm, with only the MDA+snail control arm within Unguja showing trends towards reduced diversity and altered inbreeding over time. The greatest differences overall, both in terms of parasite fecundity and genetic sub-structuring, were observed between the islands, consistent with Pemba's persistently higher mean infection intensities compared to neighbouring Unguja, and within islands in terms of infection hotspots (across three definitions). These findings highlight the important contribution of population genetic analyses to elucidate extensive genetic diversity and biological drivers, including potential gene-environmental factors, that may override short term selective pressures imposed by differential disease control strategies. Trial Registration: ClinicalTrials.gov ISRCTN48837681.

Diketopyrrolo[3,4-c]pyrrole derivative as a promising ligand for the stabilization of G-quadruplex DNA structures.

Telomerase, oncogenes and tumor suppressors are closely associated with tumour occurrence, therefore these structures are being recognized as targets for the development of new anticancer drugs. The efficacy of several molecules in telomerase inhibition and regulation of genes expression, by adduct formation with G-quadruplexes (G4), has been studied by biophysical and biochemical methods with promising results. We report here the synthesis and structural characterization of a small positively charged diketopyrrolo[3,4-c]pyrrole derivative, identified as DPP(PyMe)2, that showed very promising results as G4 stabilizing ligand. The data obtained from UV-Vis and fluorescence experiments suggest that DPP(PyMe)2 presents high affinity to G4 structures. Docking studies and molecular dynamics simulations unraveled the binding modes of the ligand with four G4 structures. The obtained results also allowed us to conclude that the DPP(PyMe)2 ligand binds into the top G-tetrad or in a mixed binding mode depending on the GQ structure. A remarkable selectivity of DPP(PyMe)2 for c-MYC and KRAS 32R in the presence of ds26 was observed by circular dichroism (CD) and fluorescence resonance energy transfer (FRET) melting experiments. CD titrations revealed a stabilization higher than 30 °C in the case of c-MYC G4 structure and, for the same sequence, DPP(PyMe)2 showed the ability to block the activity of Taq polymerase in a dose-dependent manner. The subcellular localization obtained with confocal microscopy corroborates the results obtained by the other techniques and the obtained data suggest that DPP(PyMe)2 is an attractive ligand for the development of G4 labelling probes.

Unraveling the Photodynamic Activity of Cationic Benzoporphyrin-Based Photosensitizers against Bladder Cancer Cells.

In this study, we report the preparation of new mono-charged benzoporphyrin complexes by reaction of the appropriate neutral benzoporphyrin with (2,2'-bipyridine)dichloroplatinum(II) and of the analogs' derivatives synthesized through alkylation of the neutral scaffold with iodomethane. All derivatives were incorporated into polyvinylpyrrolidone (PVP) micelles. The ability of the resultant formulations to generate reactive oxygen species was evaluated, mainly the singlet oxygen formation. Then, the capability of the PVP formulations to act as photosensitizers against bladder cancer cells was assessed. Some of the studied formulations were the most active photosensitizers causing a decrease in HT-1376 cells' viability. This creates an avenue to further studies related to bladder cancer cells.

Antimicrobial Photodynamic Approach in the Inactivation of Viruses in Wastewater: Influence of Alternative Adjuvants.

Pathogenic viruses are frequently present in marine and estuarine waters, due to poor wastewater (WW) treatments, which consequently affect water quality and human health. Chlorination, one of the most common methods used to ensure microbiological safety in tertiarily treated effluents, may lead to the formation of toxic chemical disinfection by-products on reaction with organic matter present in the effluents. Antimicrobial photodynamic therapy (aPDT) can be a promising disinfecting approach for the inactivation of pathogens, without the formation of known toxic by-products. Additionally, some studies have reported the potentiator effect on aPDT of some compounds, such as potassium iodide (KI) and hydrogen peroxide (H2O2). In the present study, the aPDT efficiency of a PS formulation constituted of five cationic porphyrins (Form) in the inactivation of E. coli T4-like bacteriophage, a model of mammalian viruses, in different aqueous matrices with different organic matter content, was evaluated. Photoinactivation studies were performed at different concentrations of Form and in the presence of the adjuvants KI and H2O2. The results showed that the efficiency of bacteriophage photoinactivation is correlated with the Form concentration, the amount of the organic matter in WW, and the adjuvant type. Form can be an effective alternative to controlling viruses in WW, particularly if combined with H2O2, allowing to significantly reduce PS concentration and treatment time. When combined with KI, the Form is less effective in inactivating T4-like bacteriophage in WW.

Revisiting density-dependent fecundity in schistosomes using sibship reconstruction.

The stability of parasite populations is regulated by density-dependent processes occurring at different stages of their life cycle. In dioecious helminth infections, density-dependent fecundity is one such regulatory process that describes the reduction in egg production by female worms in high worm burden within-host environments. In human schistosomiasis, the operation of density-dependent fecundity is equivocal and investigation is hampered by the inaccessibility of adult worms that are located intravascularly. Current understanding is almost exclusively limited to data collected from two human autopsy studies conducted over 40 years ago, with subsequent analyses having reached conflicting conclusions. Whether egg production is regulated in a density-dependent manner is key to predicting the effectiveness of interventions targeting the elimination of schistosomiasis and to the interpretation of parasitological data collected during monitoring and evaluation activities. Here, we revisit density-dependent fecundity in the two most globally important human Schistosoma spp. using a statistical modelling approach that combines molecular inference on the number of parents/adult worms in individual human hosts with parasitological egg count data from mainland Tanzania and Zanzibar. We find a non-proportional relationship between S. haematobium egg counts and inferred numbers of female worms, providing the first clear evidence of density-dependent fecundity in this schistosome species. We do not find robust evidence for density-dependent fecundity in S. mansoni because of high sensitivity to some modelling assumptions and the lower statistical power of the available data. We discuss the strengths and limitations of our model-based analytical approach and its potential for improving our understanding of density dependence in schistosomiasis and other human helminthiases earmarked for elimination.

Breast cancer in very young women-a multicenter 10-year experience.

BACKGROUND: Breast cancer (BC) is the most prevalent cancer in adult young women in Europe. Although rare, it is one of the leading causes of death in this age group. The aim of this study is to characterize a cohort of young women regarding tumor stage, biology, treatment and survival. PATIENTS AND METHODS: We present a multicenter retrospective analysis of women <35 years of age, diagnosed with BC between 2008 and 2017. A total of 207 patients from five Portuguese centers were included, from whom 172 were eligible for analysis. Data were analyzed using IBM SPPSS statistics. RESULTS: Median age at diagnosis was 31 years. Fifty-one percent of tumors were hormone receptor (HR)-positive/human epidermal growth factor receptor 2 (HER2)-negative, 20% HR-positive/HER2-positive, 8% HR-negative/HER2-positive and 20% triple-negative BC. Twenty-two percent of patients were diagnosed in stage I, 26% stage II, 45% stage III and 6% had de novo metastatic cancer. Thirty-nine percent of patients were treated with neoadjuvant chemotherapy. Mean follow-up time was 64.9 months and overall survival at 5 years, of the entire cohort and metastatic patients, was 86.5% and 26%, respectively. CONCLUSIONS: In our study we found similar population characteristics to other cohorts <35 years of age. To our knowledge, this is one of the largest cohorts in very young women. BC in young women is an important issue and further studies are needed to provide better care and survivorship to patients.

Performance assessment of 11 commercial serological tests for SARS-CoV-2 on hospitalised COVID-19 patients.

BACKGROUND: Commercial availability of serological tests to evaluate immunoglobulins (Ig) targeting severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) has grown exponentially since the start of the coronavirus disease 2019 (COVID-19) outbreak. Thorough validation of these tests is important before use as epidemiological tools to infer seroprevalence in specific populations and as diagnostic tools to complement molecular approaches (e.g., quantitative reverse transcription-polymerase chain reaction). METHODS: Commercial serological tests from 11 suppliers were assayed side-by-side using 126 samples from SARS-CoV-2-infected inpatients and 36 from healthy and HIV-infected individuals. RESULTS: The majority of the tests assayed have >95% specificity. For the sensitivity calculation, samples were stratified by days since symptoms onset; sensitivity peaks at 16-21 days for IgM and IgA (maximum 91.2%, Euroimmun) and, dependant on the test, at 16-21 or >21 days for IgG (maximum 94.1%, Snibe). Data from semiquantitative tests show that patients with a severe clinical presentation have lower levels of Ig targeting SARS-CoV-2 at <10 days since symptoms onset and higher levels at >21 days, compared to patients with a non-severe presentation. CONCLUSIONS: This study highlights the heterogeneity of sensitivity and generally high specificity of the serological tests and establishes a basis for their usefulness to complement diagnostic techniques and population seroprevalence studies.

Evaluation of the cellular protection by novel spiropyrazole compounds in dopaminergic cell death.

The loss of neurons is strongly correlated with aging and aging-associated disorders. In this study, cell viability assays and mitochondrial function were performed to evaluate the effect of new spiro-pyrazole derivatives, prepared from aldehydes and 3-amino-1-phenyl-2-pyrazolin-5-one, on neuroprotection in an in vitro model of dopaminergic cell death induced by 1-methyl-4-phenylpyridinium (MPP+). The percentages of neuroprotection by derivatives were found between 21.26% and 52.67% at selected concentrations (10-50 µM) with compound 4d exerting the best neuroprotective effect. The results show that the studied spiropyrazolones perform important roles in dopaminergic neuroprotection and can be used for potential new therapies in the treatment of neurodegenerative disorders including Parkinson's disease.