Intracranial non-germinomatous germ cell tumors in children and adolescents: how can the experience from an uppermiddle-income country contribute to the worldwide effort to improve outcomes?

Background: Non-germinomatous germ cell tumors (NGGCT) accounts for one third of intracranial GCT. While the germinoma group have an excellent overall survival, the standard of practice for children with NGGCT is still under evaluation. Aims: Describe the results of the of the Brazilian consortium protocol. Methods: Since 2013, 15 patients with a diagnosis of NGGCT by histopathology and/or serum/cerebrospinal fluid (CSF) tumor markers, ßHCG >200mlU/ml and/or positive alpha-fetoprotein were treated with neoadjuvant chemotherapy with carboplatin, cyclophosphamide and etoposide followed by ventricular radiotherapy (RTV) of 18Gy with boost (32Gy) to the primary site. Metastatic patients underwent craniospinal irradiation (CSI) and "slow responders" to the four initial cycles of CT, to autologous stem cell transplantation (ASCT) followed by CSI. Results: Mean age, 13.1 years. Thirteen males. Primary sites: pineal (n=12), suprasellar (n=2) and bifocal (n=1). Four patients were metastatic at diagnosis. Eight patients had CSF and/or serum alpha-fetoprotein levels > 1,000ng/ml. Tumor responses after chemotherapy demonstrated complete in six cases and partial in seven, with "second-look" surgery being performed in five cases, and two patients presenting viable lesions being referred to ASCT. The main toxicity observed was hematological grades 3/4. Two patients with metastatic disease, one with Down Syndrome and AFP > 1,000ng/ml and the other with choriocarcinoma and pulmonary metastases, developed progressive disease resulting in death, as well as two other patients without evidence of disease, due to endocrinological disorders. Event-free and overall survival at 2 and 5 years were 80% and 72.7%, respectively, with a mean follow-up of 48 months (range, 7-107). Conclusions: Despite the small number of patients, in our series, treatment with six cycles of chemotherapy and RTV with focal boost for localized disease (n=11) and ACST for identified slow responders (n=2) seem to be effective strategies contributing to the overall effort to improve outcomes of this group of patients.

Chitosan/Virgin Coconut Oil-Based Emulsions Doped with Photosensitive Curcumin Loaded Capsules: A Functional Carrier to Topical Treatment.

In recent years, there has been a growing interest in developing smart drug delivery systems based on natural resources combined with stimulus-sensitive elements. This trend aims to formulate innovative and sustainable delivery platforms tailored for topical applications. This work proposed the use of layer-by-layer (LbL) methodology to fabricate biocompatible photo-responsive multilayer systems. These systems are composed of a polyoxometalate inorganic salt (POM) ([NaP5W30O110]14-) and a natural origin polymer, chitosan (CHT). Curcumin (CUR), a natural bioactive compound, was incorporated to enhance the functionality of these systems during the formation of hollow capsules. The capsules produced, with sizes between 2-5µm (SEM), were further dispersed into CHT/VCO (virgin coconut oil) emulsion solutions that were casted into molds and dried at 37 °C for 48 h. The system presented a higher water uptake in PBS than in acidic conditions, still significantly lower than that earlier reported to other CHT/VCO-based systems. The drug release profile is not significantly influenced by the medium pH reaching a maximum of 37% ± 1% after 48 h. The antioxidant performance of the designed structures was further studied, suggesting a synergistic beneficial effect resulting from CUR, POM, and VCO individual bioactivities. The increased amount of those excipients released to the media over time promoted an increase in the antioxidant activity of the system, reaching a maximum of 38.1% ± 0.1% after 48 h. This work represents a promising step towards developing advanced, sustainable drug delivery systems for topical applications.

Human osteoclasts in vitro are dose dependently both inhibited and stimulated by cannabidiol (CBD) and Δ9-tetrahydrocannabinol (THC).

Legalized use of cannabis for medical or recreational use is becoming more and more common. With respect to potential side-effects on bone health only few clinical trials have been conducted - and with opposing results. Therefore, it seems that there is a need for more knowledge on the potential effects of cannabinoids on human bone cells. We studied the effect of cannabidiol (CBD) and Δ9-tetrahydrocannabinol (THC) (dose range from 0.3 to 30 µM) on human osteoclasts in mono- as well as in co-cultures with human osteoblast lineage cells. We have used CD14+ monocytes from anonymous blood donors to differentiate into osteoclasts, and human osteoblast lineage cells from outgrowths of human trabecular bone. Our results show that THC and CBD have dose-dependent effects on both human osteoclast fusion and bone resorption. In the lower dose ranges of THC and CBD, osteoclast fusion was unaffected while bone resorption was increased. At higher doses, both osteoclast fusion and bone resorption were inhibited. In co-cultures, both osteoclastic bone resorption and alkaline phosphatase activity of the osteoblast lineage cells were inhibited. Finally, we observed that the cannabinoid receptor CNR2 is more highly expressed than CNR1 in CD14+ monocytes and pre-osteoclasts, but also that differentiation to osteoclasts was coupled to a reduced expression of CNR2, in particular. Interestingly, under co-culture conditions, we only detected the expression of CNR2 but not CNR1 for both osteoclast as well as osteoblast lineage nuclei. In line with the existing literature on the effect of cannabinoids on bone cells, our current study shows both stimulatory and inhibitory effects. This highlights that potential unfavorable effects of cannabinoids on bone cells and bone health is a complex matter. The contradictory and lacking documentation for such potential unfavorable effects on bone health as well as other potential effects, should be taken into consideration when considering the use of cannabinoids for both medical and recreational use.

FLT3 tyrosine kinase inhibition modulates PRC2 and promotes differentiation in acute myeloid leukemia.

Internal tandem duplication mutations in fms-like tyrosine kinase 3 (FLT3-ITD) are recurrent in acute myeloid leukemia (AML) and increase the risk of relapse. Clinical responses to FLT3 inhibitors (FLT3i) include myeloid differentiation of the FLT3-ITD clone in nearly half of patients through an unknown mechanism. We identified enhancer of zeste homolog 2 (EZH2), a component of polycomb repressive complex 2 (PRC2), as a mediator of this effect using a proteomic-based screen. FLT3i downregulated EZH2 protein expression and PRC2 activity on H3K27me3. FLT3-ITD and loss-of-function mutations in EZH2 are mutually exclusive in human AML. We demonstrated that FLT3i increase myeloid maturation with reduced stem/progenitor cell populations in murine Flt3-ITD AML. Combining EZH1/2 inhibitors with FLT3i increased terminal maturation of leukemic cells and reduced leukemic burden. Our data suggest that reduced EZH2 activity following FLT3 inhibition promotes myeloid differentiation of FLT3-ITD leukemic cells, providing a mechanistic explanation for the clinical observations. These results demonstrate that in addition to its known cell survival and proliferation signaling, FLT3-ITD has a second, previously undefined function to maintain a myeloid stem/progenitor cell state through modulation of PRC2 activity. Our findings support exploring EZH1/2 inhibitors as therapy for FLT3-ITD AML.

Oropharyngeal colostrum immunotherapy and risk reduction of mortality in very low birth weight premature newborns: a clinical trial.

OBJECTIVE: To evaluate the effect of oropharyngeal colostrum immunotherapy (OCI) on the mortality of preterm newborns (PTNB) with very low birth weight (VLBW). METHOD: Non-randomized clinical trial, carried out with 138 mother-child pairs attended at a public maternity hospital. The treatment group used raw colostrum, dripping 4 drops (0.2 ml) into the oropharyngeal mucosa, totaling 8 administrations in 24 h, up to the 7th complete day of life (OCI). The control group was composed of newborns admitted to the same maternity hospital before the implementation of the OCI. Analyzes were performed: descriptive, bivariate, multiple logistic regression, and survival analysis, with a significance level of 5% and 95% CI. RESULTS: The treatment group had an RR of death of 0.26 (95% CI = 0.07-0.67; p = 0.00), adjusted for maternal age, marital status, gestational hypertension, type of delivery, number of prenatal visits, and birth weight. Number Needed to Treat (NNT) demonstrated that for every 5 individuals treated with OCI, one death was prevented NNT = 4.9 (95% CI = 1.84-5.20); however, for PTNB with VLBW who remained hospitalized for 50, 100 and 150 days, the NNT reduces to 4, 4 and 3, respectively. CONCLUSION: The OCI proved to be a beneficial intervention, since it reduced the risk of mortality in PTNB with VLBW when compared to the control group.

3D bioactive ionic liquid-based architectures: An anti-inflammatory approach for early-stage osteoarthritis.

3D bioprinting enables the fabrication of biomimetic cell-laden constructs for cartilage regeneration, offering exclusive strategies for precise pharmacological screenings in osteoarthritis (OA). Synovial inflammation plays a crucial role in OA's early stage and progression, characterized by the increased of the synovial pro-inflammatory mediators and cytokines and chondrocyte apoptosis. Therefore, there is an urgent need to develop solutions for effectively managing the primary events associated with OA. To address these issues, a phenolic-based biocompatible ionic liquid approach, combining alginate (ALG), acemannan (ACE), and cholinium caffeate (Ch[Caffeate]), was used to produce easily printable bioinks. Through the use of this strategy 3D constructs with good printing resolution and high structural integrity were obtained. The encapsulation of chondrocytes like ATDC5 cells provided structures with good cell distribution, viability, and growth, for up to 14 days. The co-culture of the constructs with THP-1 macrophages proved their ability to block pro-inflammatory cytokines (TNF-α and IL-6) and mediators (GM-CSF), released by the cultured cells. Moreover, incorporating the biocompatible ionic liquid into the system significantly improved its bioactive performance without compromising its physicochemical features. These findings demonstrate that ALG/ACE/Ch[Caffeate] bioinks have great potential for bioengineering cartilage tissue analogs. Besides, the developed ALG/ACE/Ch[Caffeate] bioinks protected encapsulated chondrocyte-like cells from the effect of the inflammation, assessed by a co-culture system with THP-1 macrophages. These results support the increasing use of Bio-ILs in the biomedical field, particularly for developing 3D bioprinting-based constructs to manage inflammatory-based changes in OA. STATEMENT OF SIGNIFICANCE: Combining natural resources with active biocompatible ionic liquids (Bio-IL) for 3D printing is herein presented as an approach for the development of tools to manage inflammatory osteoarthritis (OA). We propose combining alginate (ALG), acemannan (ACE), and cholinium caffeate (Ch[Caffeate]), a phenolic-based Bio-IL with anti-inflammatory and antioxidant features, to produce bioinks that allow to obtain 3D constructs with good printing resolution, structural integrity, and that provide encapsulated chondrocyte-like cells good viability. The establishment of a co-culture system using the printed constructs and THP-1-activated macrophages allowed us to study the encapsulated chondrocyte-like cells behaviour within an inflammatory scenario, a typical event in early-stage OA. The obtained outcomes support the beneficial use of Bio-ILs in the biomedical field, particularly for the development of 3D bioprinting-based models that allow the monitoring of inflammatory-based events in OA.

Antifungal potential of the new copper(II)-theophylline/1,10-phenanthroline complex against drug-resistant Candida species.

Candida spp. are the commonest fungal pathogens worldwide. Antifungal resistance is a problem that has prompted the discovery of novel anti-Candida drugs. Herein, 25 compounds, some of them containing copper(II), cobalt(II) and manganese(II) ions, were initially evaluated for inhibiting the growth of reference strains of Candida albicans and Candida tropicalis. Eight (32%) of the compounds inhibited the proliferation of these yeasts, displaying minimum inhibitory concentrations (MICs) ranging from 31.25 to 250 µg/mL and minimum fungicidal concentration (MFCs) from 62.5 to 250 µg/mL. Drug-likeness/pharmacokinetic calculated by SwissADME indicated that the 8 selected compounds were suitable for use as topical drugs. The complex CTP, Cu(theo)2phen(H2O).5H2O (theo = theophylline; phen = 1,10-phenanthroline), was chosen for further testing against 10 medically relevant Candida species that were resistant to fluconazole/amphotericin B. CTP demonstrated a broad spectrum of action, inhibiting the growth of all 20 clinical fungal isolates, with MICs from 7.81 to 62.5 µg/mL and MFCs from 15.62 to 62.5 µg/mL. Conversely, CTP did not cause lysis in erythrocytes. The toxicity of CTP was evaluated in vivo using Galleria mellonella and Tenebrio molitor. CTP had no or low levels of toxicity at doses ranging from 31.25 to 250 µg/mL for 5 days. After 24 h of treatment, G. mellonella larvae exhibited high survival rates even when exposed to high doses of CTP (600 µg/mL), with the 50% cytotoxic concentration calculated as 776.2 µg/mL, generating selectivity indexes varying from 12.4 to 99.4 depending on each Candida species. These findings suggest that CTP could serve as a potential drug to treat infections caused by Candida species resistant to clinically available antifungals.

Oropharyngeal colostrum immunotherapy and risk reduction of mortality in very low birth weight premature newborns: a clinical trial

Abstract Objective To evaluate the effect of oropharyngeal colostrum immunotherapy (OCI) on the mortality of preterm newborns (PTNB) with very low birth weight (VLBW). Method Non-randomized clinical trial, carried out with 138 mother-child pairs attended at a public maternity hospital. The treatment group used raw colostrum, dripping 4 drops (0.2 ml) into the oropharyngeal mucosa, totaling 8 administrations in 24 h, up to the 7th complete day of life (OCI). The control group was composed of newborns admitted to the same maternity hospital before the implementation of the OCI. Analyzes were performed: descriptive, bivariate, multiple logistic regression, and survival analysis, with a significance level of 5% and 95% CI. Results The treatment group had an RR of death of 0.26 (95% CI = 0.07-0.67; p= 0.00), adjusted for maternal age, marital status, gestational hypertension, type of delivery, number of prenatal visits, and birth weight. Number Needed to Treat (NNT) demonstrated that for every 5 individuals treated with OCI, one death was prevented NNT = 4.9 (95% CI = 1.84-5.20); however, for PTNB with VLBW who remained hospitalized for 50, 100 and 150 days, the NNT reduces to 4, 4 and 3, respectively. Conclusion The OCI proved to be a beneficial intervention, since it reduced the risk of mortality in PTNB with VLBW when compared to the control group.

Development of Echinocandin Resistance in Candida haemulonii: An Emergent, Widespread, and Opportunistic Fungal Pathogen.

Echinocandins, used for the prevention and treatment of invasive fungal infections, have led to a rise in breakthrough infections caused by resistant Candida species. Among these species, those belonging to the Candida haemulonii complex are rare multidrug-resistant (MDR) yeasts that are frequently misidentified but have emerged as significant healthcare-associated pathogens causing invasive infections. The objectives of this study were to investigate the evolutionary pathways of echinocandin resistance in C. haemulonii by identifying mutations in the FKS1 gene and evaluating the impact of resistance on fitness. After subjecting a MDR clinical isolate of C. haemulonii (named Ch4) to direct selection using increasing caspofungin concentrations, we successfully obtained an isolate (designated Ch4'r) that exhibited a high level of resistance, with MIC values exceeding 16 mg/L for all tested echinocandin drugs (caspofungin, micafungin, and anidulafungin). Sequence analysis revealed a specific mutation in the resistant Ch4'r strain, leading to an arginine-histidine amino acid substitution (R1354H), occurring at the G4061A position of the HS2 region of the FKS1 gene. Compared to the wild-type strain, Ch4'r exhibited significantly reduced growth proliferation, biofilm formation capability, and phagocytosis ratio, indicating a decrease in fitness. Transmission electron microscopy analysis revealed alterations in cell wall components, with a notable increase in cell wall thickness. The resistant strain also exhibited higher amounts (2.5-fold) of chitin, a cell wall-located molecule, compared to the wild-type strain. Furthermore, the resistant strain demonstrated attenuated virulence in the Galleria mellonella larval model. The evolved strain Ch4'r maintained its resistance profile in vivo since the treatment with either caspofungin or micafungin did not improve larval survival or reduce the fungal load. Taken together, our findings suggest that the acquisition of pan-echinocandin resistance occurred rapidly after drug exposure and was associated with a significant fitness cost in C. haemulonii. This is particularly concerning as echinocandins are often the first-line treatment option for MDR Candida species.

Chagas Disease Control-Many Approaches to Prospect.

Chagas disease is an emerging and neglected tropical disease caused by the protozoan parasite Trypanosoma cruzi, estimated to infect 8 to 10 million people worldwide, according to the World Health Organization [...].

Chitosan/Virgin-Coconut-Oil-Based System Enriched with Cubosomes: A 3D Drug-Delivery Approach.

Emulsion-based systems that combine natural polymers with vegetable oils have been identified as a promising research avenue for developing structures with potential for biomedical applications. Herein, chitosan (CHT), a natural polymer, and virgin coconut oil (VCO), a resource obtained from coconut kernels, were combined to create an emulsion system. Phytantriol-based cubosomes encapsulating sodium diclofenac, an anti-inflammatory drug, were further dispersed into CHT/VCO- based emulsion. Then, the emulsions were frozen and freeze-dried to produce scaffolds. The scaffolds had a porous structure ranging from 20.4 to 73.4 µm, a high swelling ability (up to 900%) in PBS, and adequate stiffness, notably in the presence of cubosomes. Moreover, a well-sustained release of the entrapped diclofenac in the cubosomes into the CHT/VCO-based system, with an accumulated release of 45 ± 2%, was confirmed in PBS, compared to free diclofenac dispersed (80 ± 4%) into CHT/VCO-based structures. Overall, the present approach opens up new avenues for designing porous biomaterials for drug delivery through a sustainable pathway.

Silver(I) and Copper(II) 1,10-Phenanthroline-5,6-dione Complexes as Promising Antivirulence Strategy against Leishmania: Focus on Gp63 (Leishmanolysin).

Leishmaniasis, caused by protozoa of the genus Leishmania, encompasses a group of neglected diseases with diverse clinical and epidemiological manifestations that can be fatal if not adequately and promptly managed/treated. The current chemotherapy options for this disease are expensive, require invasive administration and often lead to severe side effects. In this regard, our research group has previously reported the potent anti-Leishmania activity of two coordination compounds (complexes) derived from 1,10-phenanthroline-5,6-dione (phendione): [Cu(phendione)3].(ClO4)2.4H2O and [Ag(phendione)2].ClO4. The present study aimed to evaluate the effects of these complexes on leishmanolysin (gp63), a virulence factor produced by all Leishmania species that plays multiple functions and is recognized as a potential target for antiparasitic drugs. The results showed that both Ag-phendione (-74.82 kcal/mol) and Cu-phendione (-68.16 kcal/mol) were capable of interacting with the amino acids comprising the active site of the gp63 protein, exhibiting more favorable interaction energies compared to phendione alone (-39.75 kcal/mol) or 1,10-phenanthroline (-45.83 kcal/mol; a classical gp63 inhibitor) as judged by molecular docking assay. The analysis of kinetic parameters using the fluorogenic substrate Z-Phe-Arg-AMC indicated Vmax and apparent Km values of 0.064 µM/s and 14.18 µM, respectively, for the released gp63. The effects of both complexes on gp63 proteolytic activity were consistent with the in silico assay, where Ag-phendione exhibited the highest gp63 inhibition capacity against gp63, with an IC50 value of 2.16 µM and the lowest inhibitory constant value (Ki = 5.13 µM), followed by Cu-phendione (IC50 = 163 µM and Ki = 27.05 µM). Notably, pretreatment of live L. amazonensis promastigotes with the complexes resulted in a significant reduction in the expression of gp63 protein, including the isoforms located on the parasite cell surface. Both complexes markedly decreased the in vitro association indexes between L. amazonensis promastigotes and THP-1 human macrophages; however, this effect was reversed by the addition of soluble gp63 molecules to the interaction medium. Collectively, our findings highlight the potential use of these potent complexes in antivirulence therapy against Leishmania, offering new insights for the development of effective treatments for leishmaniasis.

Building Fucoidan/Agarose-Based Hydrogels as a Platform for the Development of Therapeutic Approaches against Diabetes.

Current management for diabetes has stimulated the development of versatile 3D-based hydrogels as in vitro platforms for insulin release and as support for the encapsulation of pancreatic cells and islets of Langerhans. This work aimed to create agarose/fucoidan hydrogels to encapsulate pancreatic cells as a potential biomaterial for diabetes therapeutics. The hydrogels were produced by combining fucoidan (Fu) and agarose (Aga), marine polysaccharides derived from the cell wall of brown and red seaweeds, respectively, and a thermal gelation process. The agarose/fucoidan (AgaFu) blended hydrogels were obtained by dissolving Aga in 3 or 5 wt % Fu aqueous solutions to obtain different proportions (4:10; 5:10, and 7:10 wt). The rheological tests on hydrogels revealed a non-Newtonian and viscoelastic behavior, while the characterization confirmed the presence of the two polymers in the structure of the hydrogels. In addition, the mechanical behavior showed that increasing Aga concentrations resulted in hydrogels with higher Young's modulus. Further, the ability of the developed materials to sustain the viability of human pancreatic cells was assessed by encapsulation of the 1.1B4HP cell line for up to 7 days. The biological assessment of the hydrogels revealed that cultured pancreatic beta cells tended to self-organize and form pseudo-islets during the period studied.

Alginate/acemannan-based beads loaded with a biocompatible ionic liquid as a bioactive delivery system.

Combining biomacromolecules with green chemistry principles and clean technologies has proven to be an effective approach for drug delivery, providing a prolonged and sustained release of the encapsulated material. The current study investigates the potential of cholinium caffeate (Ch[Caffeate]), a phenolic-based biocompatible ionic liquid (Bio-IL) entrapped in alginate/acemannan beads, as a drug delivery system able to reduce local joint inflammation on osteoarthritis (OA) treatment. The synthesized Bio-IL has antioxidant and anti-inflammatory actions that, combined with biopolymers as 3D architectures, promote the entrapment and sustainable release of the bioactive molecules over time. The physicochemical and morphological characterization of the beads (ALC, ALAC0,5, ALAC1, and ALAC3, containing 0, 0.5, 1, and 3 %(w/v) of Ch[Caffeate], respectively) revealed a porous and interconnected structure, with medium pore sizes ranging from 209.16 to 221.30 µm, with a high swelling ability (up 2400 %). Ch[Caffeate] significantly improved the antioxidant activities of the constructs by 95 % and 97 % for ALAC1 and ALAC3, respectively, when compared to ALA (56 %). Besides, the structures provided the environment for ATDC5 cell proliferation, and cartilage-like ECM formation, supported by the increased GAGs in ALAC1 and ALAC3 formulations after 21 days. Further, the ability to block the secretion of pro-inflammatory cytokines (TNF-α and IL-6), from differentiated THP-1 was evidenced by ChAL-Ch[Caffeate] beads. These outcomes suggest that the established strategy based on using natural and bioactive macromolecules to develop 3D constructs has great potential to be used as therapeutic tools for patients with OA.

Outcome of Children and Adolescents With Primary Intracranial Germinoma Treated With Chemotherapy and Reduced Dose-Field Irradiation: A Prospective Brazilian Experience.

PURPOSE: This prospective Brazilian single-arm trial was conducted to determine response to chemotherapy and survival after response-based radiotherapy in children with intracranial germinomas, in the setting of a multi-institutional study in a middle-income country (MIC) with significant disparity of subspecialty care. PATIENTS AND METHODS: Since 2013, 58 patients with histologic and/or serum and CSF tumor marker evaluations of primary intracranial germ cell tumors were diagnosed; 43 were germinoma with HCGß levels ≤200 mIU/mL and five between 100 and 200 mIU/mL. The treatment plan consisted of four cycles of carboplatin and etoposide followed by 18 Gy whole-ventricular field irradiation (WVFI) and primary site(s) boost up to 30 Gy; 24 Gy craniospinal was prescribed for disseminated disease. RESULTS: Mean age 13.2 years (range, 4.7-25.5 years); 29 were males. Diagnosis was made by tumor markers (n = 6), surgery (n = 25), or both (n = 10). Two bifocal cases with negative tumor markers were treated as germinoma. Primary tumor location was pineal (n = 18), suprasellar (n = 14), bifocal (n = 10), and basal ganglia/thalamus (n = 1). Fourteen had ventricular/spinal spread documented by imaging studies. Second-look surgery occurred in three patients after chemotherapy. Thirty-five patients achieved complete responses after chemotherapy, and eight showed residual teratoma/scar. Toxicity was mostly grade 3/4 neutropenia/thrombocytopenia during chemotherapy. At a median follow-up of 44.5 months, overall and event-free survivals were 100%. CONCLUSION: The treatment is tolerable, and WVFI dose reduction to 18 Gy preserves efficacy; we have demonstrated the feasibility of successfully conducting a prospective multicenter trial in a large MIC despite resource disparity.

Short- and long-term follow-up after fecal microbiota transplantation as treatment for recurrent Clostridioides difficile infection in patients with inflammatory bowel disease.

Background: Patients with inflammatory bowel disease (IBD) are at an increased risk of developing Clostridioides difficile infection (CDI). Treatment of CDI in patients with IBD is challenging due to higher failure rates and concomitant IBD activity. Objectives: We performed a multicentre cohort study in patients with IBD who received fecal microbiota transplantation (FMT) for recurrent CDI (rCDI), to further investigate factors that influence the clinical outcome and course of both rCDI and IBD. Design: This is a multicentre cohort study conducted in five European FMT centres. Methods: Adult IBD patients treated with FMT for rCDI were studied. Cure was defined as clinical resolution of diarrhoea or diarrhoea with a negative C. difficile test. The definition of an IBD flare was record based. Long-term follow-up data were collected including new episodes of CDI, IBD flares, infections, hospital admissions, and death. Results: In total, 113 IBD patients underwent FMT because of rCDI. Mean age of the patients was 48 years; 64% had ulcerative colitis. Concomitant rCDI was associated with an IBD flare in 54%, of whom 63% had received IBD remission-induction therapy prior to FMT. All FMT procedures were preceded by vancomycin treatment, 40% of patients received FMT via colonoscopy. CDI cure rate was 71%. Long-term follow-up data were available in 90 patients with a median follow-up of 784 days (402-1251). IBD activity decreased in 39% of patients who had active IBD at baseline, whereas an IBD flare occurred in only 5%. During follow-up of up to 2 years, 27% of the patients had infections, 39% were hospitalized, 5% underwent colectomy, and 10% died (median age of these latter patients: 72 years). Conclusion: FMT for rCDI in IBD patients is safe and effective, and IBD exacerbation after FMT is infrequent. Further studies should investigate the effects on IBD course following FMT.

SARS-CoV-2 Post-Infection and Sepsis by Saccharomyces cerevisiae: A Fatal Case Report-Focus on Fungal Susceptibility and Potential Virulence Attributes.

The pandemic caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has been responsible for approximately 6.8 million deaths worldwide, threatening more than 753 million individuals. People with severe coronavirus disease-2019 (COVID-19) infection often exhibit an immunosuppression condition, resulting in greater chances of developing co-infections with bacteria and fungi, including opportunistic yeasts belonging to the Saccharomyces and Candida genera. In the present work, we have reported the case of a 75-year-old woman admitted at a Brazilian university hospital with an arterial ulcer in the left foot, which was being prepared for surgical amputation. The patient presented other underlying diseases and presented positive tests for COVID-19 prior to hospitalization. She received antimicrobial treatment, but her general condition worsened quickly, leading to death by septic shock after 4 days of hospitalization. Blood samples collected on the day she died were positive for yeast-like organisms, which were later identified as Saccharomyces cerevisiae by both biochemical and molecular methods. The fungal strain exhibited low minimal inhibitory concentration values for the antifungal agents tested (amphotericin B, 5-flucytosine, caspofungin, fluconazole and voriconazole), and it was able to produce important virulence factors, such as extracellular bioactive molecules (e.g., aspartic peptidase, phospholipase, esterase, phytase, catalase, hemolysin and siderophore) and biofilm. Despite the activity against planktonic cells, the antifungals were not able to impact the mature biofilm parameters (biomass and viability). Additionally, the S. cerevisiae strain caused the death of Tenebrio molitor larvae, depending on the fungal inoculum, and larvae immunosuppression with corticosteroids increased the larvae mortality rate. In conclusion, the present study highlighted the emergence of S. cerevisiae as an opportunistic fungal pathogen in immunosuppressed patients presenting several severe comorbidities, including COVID-19 infection.

Apicobasal transferrin receptor localization and trafficking in brain capillary endothelial cells.

The detailed mechanisms by which the transferrin receptor (TfR) and associated ligands traffic across brain capillary endothelial cells (BECs) of the CNS-protective blood-brain barrier constitute an important knowledge gap within maintenance and regulation of brain iron homeostasis. This knowledge gap also presents a major obstacle in research aiming to develop strategies for efficient receptor-mediated drug delivery to the brain. While TfR-mediated trafficking from blood to brain have been widely studied, investigation of TfR-mediated trafficking from brain to blood has been limited. In this study we investigated TfR distribution on the apical and basal plasma membranes of BECs using expansion microscopy, enabling sufficient resolution to separate the cellular plasma membranes of these morphological flat cells, and verifying both apical and basal TfR membrane domain localization. Using immunofluorescence-based transcellular transport studies, we delineated endosomal sorting of TfR endocytosed from the apical and basal membrane, respectively, as well as bi-directional TfR transcellular transport capability. The findings indicate different intracellular sorting mechanisms of TfR, depending on the apicobasal trafficking direction across the BBB, with the highest transcytosis capacity in the brain-to-blood direction. These results are of high importance for the current understanding of brain iron homeostasis. Also, the high level of TfR trafficking from the basal to apical membrane of BECs potentially explains the low transcytosis which are observed for the TfR-targeted therapeutics to the brain parenchyma.

The Anti-Leishmania amazonensis and Anti-Leishmania chagasi Action of Copper(II) and Silver(I) 1,10-Phenanthroline-5,6-dione Coordination Compounds.

Leishmaniasis is a neglected disease caused by protozoa belonging to the Leishmania genus. Notably, the search for new, promising and potent anti-Leishmania compounds remains a major goal due to the inefficacy of the available drugs used nowadays. In the present work, we evaluated the effects of 1,10-phenanthroline-5,6-dione (phendione) coordinated to silver(I), [Ag(phendione)2]ClO4 (Ag-phendione), and copper(II), [Cu(phendione)3](ClO4)2·4H2O (Cu-phendione), as potential drugs to be used in the chemotherapy against Leishmania amazonensis and Leishmania chagasi. The results showed that promastigotes treated with Ag-phendione and Cu-phendione presented a significant reduction in the proliferation rate. The IC50 values calculated to Ag-phendione and Cu-phendione, respectively, were 7.8 nM and 7.5 nM for L. amazonensis and 24.5 nM and 20.0 nM for L. chagasi. Microscopical analyses revealed several relevant morphological changes in promastigotes, such as a rounding of the cell body and a shortening/loss of the single flagellum. Moreover, the treatment promoted alterations in the unique mitochondrion of these parasites, inducing significant reductions on both metabolic activity and membrane potential parameters. All these cellular perturbations induced the triggering of apoptosis-like death in these parasites, as judged by the (i) increased percentage of annexin-positive/propidium iodide negative cells, (ii) augmentation in the proportion of parasites in the sub-G0/G1 phase and (iii) DNA fragmentation. Finally, the test compounds showed potent effects against intracellular amastigotes; contrarily, these molecules were well tolerated by THP-1 macrophages, which resulted in excellent selective index values. Overall, the results highlight new selective and effective drugs against Leishmania species, which are important etiological agents of both cutaneous (L. amazonensis) and visceral (L. chagasi) leishmaniasis in a global perspective.