Intestinal Permeability and IgA Provoke Immune Vasculitis Linked to Cardiovascular Inflammation.

Recent experimental data and clinical, genetic, and transcriptome evidence from patients converge to suggest a key role of interleukin-1ß (IL-1ß) in the pathogenesis of Kawasaki disease (KD). However, the molecular mechanisms involved in the development of cardiovascular lesions during KD vasculitis are still unknown. Here, we investigated intestinal barrier function in KD vasculitis and observed evidence of intestinal permeability and elevated circulating secretory immunoglobulin A (sIgA) in KD patients, as well as elevated sIgA and IgA deposition in vascular tissues in a mouse model of KD vasculitis. Targeting intestinal permeability corrected gut permeability, prevented IgA deposition and ameliorated cardiovascular pathology in the mouse model. Using genetic and pharmacologic inhibition of IL-1ß signaling, we demonstrate that IL-1ß lies upstream of disrupted intestinal barrier function, subsequent IgA vasculitis development, and cardiac inflammation. Targeting mucosal barrier dysfunction and the IL-1ß pathway may also be applicable to other IgA-related diseases, including IgA vasculitis and IgA nephropathy.

Transcriptomic responses of Mediterranean sponges upon encounter with symbiont microbial consortia.

BACKGROUND: Sponges (phylum Porifera) constantly interact with microbes. They graze on microbes from the water column by filter-feeding and they harbor symbiotic partners within their bodies. In experimental setups, sponges take up symbionts at lower rates compared with seawater microbes. This suggests that sponges have the capacity to differentiate between microbes and preferentially graze in non-symbiotic microbes, although the underlying mechanisms of discrimination are still poorly understood. Genomic studies showed that, compared to other animal groups, sponges present an extended repertoire of immune receptors, in particular NLRs, SRCRs, and GPCRs, and a handful of experiments showed that sponges regulate the expression of these receptors upon encounter with microbial elicitors. We hypothesize that sponges may rely on differential expression of their diverse repertoire of poriferan immune receptors to sense different microbial consortia while filter-feeding. To test this, we characterized the transcriptomic response of two sponge species, Aplysina aerophoba and Dysidea avara, upon incubation with microbial consortia extracted from A. aerophoba in comparison with incubation with seawater microbes. The sponges were sampled after 1 h, 3 h, and 5 h for RNA-Seq differential gene expression analysis. RESULTS: D. avara incubated with A. aerophoba-symbionts regulated the expression of genes related to immunity, ubiquitination, and signaling. Within the set of differentially-expressed immune genes we identified different families of Nucleotide Oligomerization Domain (NOD)-Like Receptors (NLRs). These results represent the first experimental evidence that different types of NLRs are involved in microbial discrimination in a sponge. In contrast, the transcriptomic response of A. aerophoba to its own symbionts involved comparatively fewer genes and lacked genes encoding for immune receptors. CONCLUSION: Our work suggests that: (i) the transcriptomic response of sponges upon microbial exposure may imply "fine-tuning" of baseline gene expression as a result of their interaction with microbes, (ii) the differential response of sponges to microbial encounters varied between the species, probably due to species-specific characteristics or related to host's traits, and (iii) immune receptors belonging to different families of NLR-like genes played a role in the differential response to microbes, whether symbionts or food bacteria. The regulation of these receptors in sponges provides further evidence of the potential role of NLRs in invertebrate host-microbe interactions. The study of sponge responses to microbes exemplifies how investigating different animal groups broadens our knowledge of the evolution of immune specificity and symbiosis.

Teaching Pathographies of Mental Illness.

This paper describes the content and evolution of a fourth-year course for medical students on teaching pathographies of mental illness. (It is a follow-up to Nathan Carlin's Pathographies of Mental Illness that appeared as an Element in the Bioethics and Neuroethics series published by Cambridge University Press.) The course originally centered on classic (and some contemporary) memoirs; however, responding to student evaluations, newer material now ensures more diversity, with material written by women and people of color, and describes the difference that can make.

Loss of kinesin-8 improves the robustness of the self-assembled spindle in Schizosaccharomyces pombe.

Chromosome segregation in female meiosis in many metazoans is mediated by acentrosomal spindles, the existence of which implies that microtubule spindles self-assemble without the participation of the centrosomes. Although it is thought that acentrosomal meiosis is not conserved in fungi, we recently reported the formation of self-assembled microtubule arrays, which were able to segregate chromosomes, in fission yeast mutants, in which the contribution of the spindle pole body (SPB; the centrosome equivalent in yeast) was specifically blocked during meiosis. Here, we demonstrate that this unexpected microtubule formation represents a bona fide type of acentrosomal spindle. Moreover, a comparative analysis of these self-assembled spindles and the canonical SPB-dependent spindle reveals similarities and differences; for example, both spindles have a similar polarity, but the location of the γ-tubulin complex differs. We also show that the robustness of self-assembled spindles can be reinforced by eliminating kinesin-8 family members, whereas kinesin-8 mutants have an adverse impact on SPB-dependent spindles. Hence, we consider that reinforced self-assembled spindles in yeast will help to clarify the molecular mechanisms behind acentrosomal meiosis, a crucial step towards better understanding gametogenesis.

Lamb-Shaffer syndrome: 20 Spanish patients and literature review expands the view of neurodevelopmental disorders caused by SOX5 haploinsufficiency.

Lamb-Shaffer Syndrome (LSS; OMIM #616803; ORPHA #313892; ORPHA #313884) is an infrequent genetic disorder that affects multiple aspects of human development especially those related to the development of the nervous system. LSS is caused by variants in the SOX5 gene. At the molecular level, SOX5 gene encodes for a transcription factor containing a High Mobility Group (HMG) DNA-Binding domain with relevant functions in brain development in different vertebrate species. Clinical features of Lamb-Shaffer syndrome may include intellectual disability, delayed speech and language development, attention deficits, hyperactivity, autism spectrum disorder, visual problems and seizures. Additionally, patients with the syndrome may present distinct facial dimorphism such as a wide mouth with full lips, small chin, broad nasal bridge, and deep-set eyes. Other physical features that have been reported in some patients include short stature, scoliosis, and joint hypermobility. Here, we report the clinical and molecular characterization of a Spanish LSS cohort of new 20 patients and review all the patients published so far which amount for 111 patients. The most frequent features included developmental delay, intellectual disability, visual problems, poor speech development and facial dysmorphic features. Strikingly, pain insensitivity and hypermetropia seems to be more frequent than previously reported, based on the frequency seen in the Spanish cohort. Eighty-three variants have been reported so far, single nucleotide variants (SNV) and copy number variants represent 47% and 53%, respectively, from the total of variants reported. Similarly to previous reports, the majority of the SNVs variants of the novel patients reported herein fall in the HMG domain of the protein. However, new variants, affecting other functional domains, were also detected. In conclusion, LLS is a rare genetic disorder mostly characterized by a wide range of developmental and neurological symptoms. Early diagnosis would allow to start of care programs, clinical follow up, prospective studies and appropriate genetic counseling, to promote clinical and social improvement to have profound lifelong benefits for patients and their families. Further research is needed to better understand the underlying mechanisms of the syndrome related to SOX5 haploinsufficiency.

NLRP3 Inflammasome Mediates Immune-Stromal Interactions in Vasculitis.

[Figure: see text].

Anthropometric characteristics of young elite sailors based on performance level.

Background: /Objectives: The aim of the present study was to analyse possible differences in anthropometric characteristics of elite sailors based on categories and performance level. Methods: ː A total of 42 young (aged 12-18 years) elite sailors (men = 31; women = 11) of the Monohull (n = 21) and Windsurfing (n = 21) categories composed the study sample. Testing was per-formed in one session the day before the start of an official and international competition. Body composition was measured using an octopolar and multi-frequency electrical bioimpedance analyser, and height was recorded using a telescopic measuring instrument. Cross-sectional study. The total sample was divided into two groups based on their performance level (ranking), 50th percentile (P1), and 100th percentile (P2). Results: ː P1 presented a lower BMI, total body fat mass, and body fat mass in the trunk, arms, and legs (p < 0.05). Similarly, P1 reported a higher total body muscle mass and body muscle mass on the trunk, arms, and legs compared to the less level performance group (p < 0.05). In addition, P2 sailors were taller and heavier (p < 0.05). Regarding categories, the Windsurf sailors presented statistically significantly lower arm fat mass than the Monohull (p < 0.05). The Windsurf sailors showed differences between the two performance-level groups (p < 0.05). Additionally, comparing the high-level performance group in both categories, higher arm muscle mass on the Windsurfing sailors was detected (p < 0.05). Conclusions: ː These findings could help to differentiate the anthropometric variables that determine sport performance in young elite sailors and could be used to differentiate the anthropometric variables in each category.

A systems-level study reveals host-targeted repurposable drugs against SARS-CoV-2 infection.

Understanding the mechanism of SARS-CoV-2 infection and identifying potential therapeutics are global imperatives. Using a quantitative systems pharmacology approach, we identified a set of repurposable and investigational drugs as potential therapeutics against COVID-19. These were deduced from the gene expression signature of SARS-CoV-2-infected A549 cells screened against Connectivity Map and prioritized by network proximity analysis with respect to disease modules in the viral-host interactome. We also identified immuno-modulating compounds aiming at suppressing hyperinflammatory responses in severe COVID-19 patients, based on the transcriptome of ACE2-overexpressing A549 cells. Experiments with Vero-E6 cells infected by SARS-CoV-2, as well as independent syncytia formation assays for probing ACE2/SARS-CoV-2 spike protein-mediated cell fusion using HEK293T and Calu-3 cells, showed that several predicted compounds had inhibitory activities. Among them, salmeterol, rottlerin, and mTOR inhibitors exhibited antiviral activities in Vero-E6 cells; imipramine, linsitinib, hexylresorcinol, ezetimibe, and brompheniramine impaired viral entry. These novel findings provide new paths for broadening the repertoire of compounds pursued as therapeutics against COVID-19.

The DNA methylation landscape of the root-knot nematode-induced pseudo-organ, the gall, in Arabidopsis, is dynamic, contrasting over time, and critically important for successful parasitism.

Root-knot nematodes (RKNs) induce giant cells (GCs) within galls which are characterized by large-scale gene repression at early stages. However, the epigenetic mechanism(s) underlying gene silencing is (are) still poorly characterized. DNA methylation in Arabidopsis galls induced by Meloidogyne javanica was studied at crucial infection stages (3 d post-infection (dpi) and 14 dpi) using enzymatic, cytological, and sequencing approaches. DNA methyltransferase mutants (met1, cmt2, cmt3, cmt2/3, drm1/2, ddc) and a DNA demethylase mutant (ros1), were analyzed for RKN resistance/tolerance, and galls were characterized by confocal microscopy and RNA-seq. Early galls were hypermethylated, and the GCs were found to be the major contributors to this hypermethylation, consistent with the very high degree of gene repression they exhibit. By contrast, medium/late galls showed no global increase in DNA methylation compared to uninfected roots, but exhibited large-scale redistribution of differentially methylated regions (DMRs). In line with these findings, it was also shown that DNA methylation and demethylation mutants showed impaired nematode reproduction and gall/GC-development. Moreover, siRNAs that were exclusively present in early galls accumulated at hypermethylated DMRs, overlapping mostly with retrotransposons in the CHG/CG contexts that might be involved in their repression, contributing to their stability/genome integrity. Promoter/gene methylation correlated with differentially expressed genes encoding proteins with basic cell functions. Both mechanisms are consistent with reprogramming host tissues for gall/GC formation. In conclusion, RNA-directed DNA methylation (RdDM; DRM2/1) pathways, maintenance methyltransferases (MET1/CMT3) and demethylation (ROS1) appear to be prominent mechanisms driving a dynamic regulation of the epigenetic landscape during RKN infection.

Brief incubation of corneal grafts in activated platelet rich plasma enhances corneal endothelial cell survival and regeneration.

Corneal transplantation is the most frequent organ transplantation worldwide. Unfortunately, corneal graft failure is common and endothelial decompensation is considered the major cause. Corneal endothelial cells (CECs) lack the capacity to reproduce, and perioperative and postoperative endothelial cell loss remains a significant challenge associated with corneal graft viability. Therefore, strategies to preserve CEC density are critical to extend graft survival. Activated platelet rich plasma (aPRP), a product extracted from autologous blood, has both antioxidant and regenerative properties. aPRP eye drops have shown effectiveness in the treatment of corneal pathologies such as ulcers, dry eye, and burns. Our purpose is to determine the protective and regenerative effect of aPRP on corneal grafts by evaluating aPRP's effect on the survival and proliferation of human CECs. Human corneal grafts were incubated in aPRP for 15 min to assess the activation of the CEC pAkt survival pathway as measured by ELISA. Evaluation of the protective effect of aPRP was made using an apoptotic model, which simulated oxidative stress conditions. Expression of apoptotic markers was measured using ELISA and endothelial cell viability was determined by optical microscopy. The CEC proliferation rate was measured in vitro with Ki-67 staining. Corneal graft gross structure was evaluated by Hematoxylin & Eosin and Masson trichrome staining. Our results indicate that a short incubation of human corneal grafts in aPRP protects CECs from apoptosis by upregulating the pAkt survival pathway and promoting CEC proliferation. Additionally, aPRP incubation does not induce histological changes in the grafts. A brief pre-treatment of human corneal grafts in aPRP may be beneficial for transplant longevity, as it protects CECs from apoptosis by upregulating intracellular survival pathways and promoting proliferation. In addition, this approach appears to be safe and has the potential to improve surgical outcomes following corneal transplantation.

The object space task shows cumulative memory expression in both mice and rats.

Declarative memory encompasses representations of specific events as well as knowledge extracted by accumulation over multiple episodes. To investigate how these different sorts of memories are created, we developed a new behavioral task in rodents. The task consists of 3 distinct conditions (stable, overlapping, and random). Rodents are exposed to multiple sample trials, in which they explore objects in specific spatial arrangements, with object identity changing from trial to trial. In the stable condition, the locations are constant during all sample trials even though the objects themselves change; in the test trial, 1 object's location is changed. In the random condition, object locations are presented in the sample phase without a specific spatial pattern. In the overlapping condition, 1 location is shared (overlapping) between all trials, while the other location changes during sample trials. We show that in the overlapping condition, instead of only remembering the last sample trial, rodents form a cumulative memory of the sample trials. Here, we could show that both mice and rats can accumulate information across multiple trials and express a long-term abstracted memory.

Simulation of the electrical stimulation of the rat brain using sleep frequencies: A finite element modeling approach.

A realistic rat brain model was used to simulate current density and electric field distributions under frequencies characteristic of sleeping states (0.8, 5, and 12 Hz). Two anode-electrode setups were simulated: plate vs. screws-anode, both with a cephalic cathode. Our simulations showed that these frequencies have limited impact on electric field and current density; however, the highest frequency evidenced higher values for both variables. The type of electrode setup had a greater effect on current distribution and induced fields. In that sense, the screws setup resulted in higher values of the modeled variables. The numeric results obtained are within the range of available data for rodent models using the finite elements method. These modeled effects should be analyzed regarding anatomical consequences (depth of penetration of the currents) and purpose of the experiment (i.e., entrainment of brain oscillations) in the context of sleep research.

Tumor necrosis factor-alpha and interferon-gamma induce inflammasome-mediated corneal endothelial cell death.

PURPOSE: Chronic corneal endothelial cell (CEC) loss results in corneal edema and vision loss in conditions such as pseudophakic bullous keratopathy (PBK), Fuchs' dystrophy, and corneal graft failure. Low CEC density has been associated with an elevation of intraocular pro-inflammatory cytokines such as tumor necrosis factor (TNF)-α and interferon (INF)-γ. These cytokines are capable of triggering pyroptosis, a programmed cell death mechanism mediated by the inflammasome, prompting the activation of the pro-inflammatory cytokine interleukin (IL)-1ß, the perpetuation of inflammation, and subsequent damage of corneal endothelial tissue. Therefore, the purpose of this study was to determine the deleterious contribution of the inflammasome and pyroptosis to CEC loss. METHODS: CECs from human donor corneas were treated ex vivo with TNF-α and IFN-γ for 48 h. Levels of caspase-1 and IL-1ß were then assayed by ELISA, and the expression of caspase-1 and gasdermin-D (GSDM-D) were confirmed by immunofluorescence. Endothelial cell damage was analyzed by a lactate dehydrogenase (LDH) release assay, and oxidative stress was determined by measuring the levels of reactive oxygen species (ROS) in the culture media. RESULTS: Inflammasome activation and oxidative stress were elevated in CECs following exposure to TNF-α and IFN-γ, which resulted in cell death by pyroptosis as determined by LDH release which was inhibited by the caspase-1 inhibitor Ac-YVAD-cmk. CONCLUSION: CEC death is induced by the pro-inflammatory cytokines TNF-α and IFN-γ, which contribute to inflammasome activation. Moreover, the inflammasome is a promising therapeutic target for the treatment of chronic CEC loss.

The Use of Biochar for Plant Pathogen Control.

To support the search for alternative, nonchemical plant disease control strategies, we present a review of the pathogen-suppressive effects of biochar, a product derived from agricultural and other organic wastes, used as a soil amendment. A wide range of biochar effects contribute to the control of root or foliar fungal pathogens through modification of root exudates, soil properties, and nutrient availability, which influence the growth of antagonist microorganisms. The induction of systemic plant defenses by biochar in the roots to reduce foliar pathogenic fungi, the activation of stress-hormone responses, as well as changes in active oxygen species are indicative of a coordinated hormonal signaling within the plant. Although scarce data are available for oomycetes and bacterial pathogens, reports indicate that biochar promotes changes in the soil microbiota influencing pathogen motility and colonization, and the induction of plant systemic defenses, both contributing to disease suppression. Biochar also suppresses nematode and insect pests. For plant-parasitic nematodes, the primary modes of action are changes in soil microbial community diversity, the release of nematicidal compounds, and the induction of plant defenses. Use of biochar-based soil amendments is a promising strategy compatible with a circular economy, based on zero waste, as part of integrated pathogen and pest management. Since biochars exert complex and distinct modes of action for the control of plant pathogens, its nature and application regimes should be designed for particular pathogens and its effects studied locally.

Clinical and electrophysiological characteristics of Guillain-Barré syndrome in Colombia.

Despite the wide literature describing the features of Guillain-Barré syndrome (GBS) in different populations worldwide, Colombian data are very scarce. We aim to characterize patients with GBS in a general hospital setting in Colombia. We conducted a retrospective chart review of GBS cases managed at the Hospital Universitario Fundación Santa Fe de Bogotá, from 2011 to 2016. Twenty-three patients were included. The most commonly reported symptoms were paresthesias (65%), pain (61%), proximal (22%) and distal (74%) limb weakness, and facial palsy (30%). 9% of patients had Fisher syndrome and 21% had other variants: Bickerstaff, pharyngeal-cervical-brachial pattern, and facial diplegia. There was a predominance of the demyelinating form (70%), with only 22% of patients presenting with the axonal variants. Our results are concordant with previous studies in Colombia.

An analysis of blood donation barriers experienced by North American and Caribbean university students in Grenada, West Indies.

OBJECTIVES: To estimate the associations of nationality, university program, donation history and gender, with blood donation barriers experienced by non-donating students on the day of a campus blood drive. This project focused particularly on nationality and the effect of the different blood donation cultures in the students' countries of origin. METHODS: A retrospective cohort study of 398 North American and Caribbean university students was conducted at St. George's University, Grenada, in 2010. Data were collected from non-donating students on campus while a blood drive was taking place. Log-binomial regression was used to estimate associations between the exposures of interest and donation barriers experienced by the students. RESULTS: North American (voluntary blood donation culture) students were more likely than Caribbean (replacement blood donation culture) students to experience "Lack of Time" (relative risk (RR) = 1.57; 95% confidence interval (CI): 1.19-2.07) and "Lack of Eligibility" (RR = 1.55; 95% CI: 1.08-2.22) as barriers to donation. Conversely, Caribbean students were a third as likely to state "Lack of Incentive" (RR = 0.32; 95% CI: 0.20-0.50), "Fear of Infection" (RR = 0.35; 95% CI: 0.21-0.58), and "Fear of Needles" (RR = 0.32; 95% CI: 0.21-0.48) were barriers than North American students. CONCLUSIONS: University students from voluntary blood donation cultures are likely to experience different barriers to donation than those from replacement cultures. Knowledge of barriers that students from contrasting blood donation systems face provides valuable information for blood drive promotion in university student populations that contain multiple nationalities.

Malignant hyperthermia-associated mutations in the S2-S3 cytoplasmic loop of type 1 ryanodine receptor calcium channel impair calcium-dependent inactivation.

Channel activities of skeletal muscle ryanodine receptor (RyR1) are activated by micromolar Ca2+ and inactivated by higher (∼1 mM) Ca2+ To gain insight into a mechanism underlying Ca2+-dependent inactivation of RyR1 and its relationship with skeletal muscle diseases, we constructed nine recombinant RyR1 mutants carrying malignant hyperthermia or centronuclear myopathy-associated mutations and determined RyR1 channel activities by [3H]ryanodine binding assay. These mutations are localized in or near the RyR1 domains which are responsible for Ca2+-dependent inactivation of RyR1. Four RyR1 mutations (F4732D, G4733E, R4736W, and R4736Q) in the cytoplasmic loop between the S2 and S3 transmembrane segments (S2-S3 loop) greatly reduced Ca2+-dependent channel inactivation. Activities of these mutant channels were suppressed at 10-100 µM Ca2+, and the suppressions were relieved by 1 mM Mg2+ The Ca2+- and Mg2+-dependent regulation of S2-S3 loop RyR1 mutants are similar to those of the cardiac isoform of RyR (RyR2) rather than wild-type RyR1. Two mutations (T4825I and H4832Y) in the S4-S5 cytoplasmic loop increased Ca2+ affinities for channel activation and decreased Ca2+ affinities for inactivation, but impairment of Ca2+-dependent inactivation was not as prominent as those of S2-S3 loop mutants. Three mutations (T4082M, S4113L, and N4120Y) in the EF-hand domain showed essentially the same Ca2+-dependent channel regulation as that of wild-type RyR1. The results suggest that nine RyR1 mutants associated with skeletal muscle diseases were differently regulated by Ca2+ and Mg2+ Four malignant hyperthermia-associated RyR1 mutations in the S2-S3 loop conferred RyR2-type Ca2+- and Mg2+-dependent channel regulation.

Two regions of the ryanodine receptor calcium channel are involved in Ca(2+)-dependent inactivation.

Skeletal (RyR1) and cardiac muscle (RyR2) isoforms of ryanodine receptor calcium channels are inhibited by millimollar Ca(2+), but the affinity of RyR2 for inhibitory Ca(2+) is ~10 times lower than that of RyR1. Previous studies demonstrated that the C-terminal quarter of RyR has critical domain(s) for Ca(2+) inactivation. To obtain further insights into the molecular basis of regulation of RyRs by Ca(2+), we constructed and expressed 18 RyR1-RyR2 chimeras in HEK293 cells and determined the Ca(2+) activation and inactivation affinities of these channels using the [(3)H]ryanodine binding assay. Replacing two distinct regions of RyR1 with corresponding RyR2 sequences reduced the affinity for Ca(2+) inactivation. The first region (RyR2 amino acids 4020-4250) contains two EF-hand Ca(2+) binding motifs (EF1, amino acids 4036-4047; EF2, amino acids 4071-4082), and the second region includes the putative second transmembrane segment (S2). A RyR1-backbone chimera containing only EF2 from RyR2 had a modest (not significant) change in Ca(2+) inactivation, whereas another chimera channel carrying only EF1 from RyR2 had a significantly reduced level of Ca(2+) inactivation. The results suggest that EF1 is a more critical determinant for RyR inactivation by Ca(2+). In addition, activities of the chimera carrying RyR2 EF-hands were suppressed at 10-100 µM Ca(2+), and the suppression was relieved by 1 mM Mg(2+). The same effects have been observed with wild-type RyR2. A mutant RyR1 carrying both regions replaced with RyR2 sequences (amino acids 4020-4250 and 4560-4618) showed a Ca(2+) inactivation affinity comparable to that of RyR2, indicating that these regions are sufficient to confer RyR2-type Ca(2+)-dependent inactivation on RyR1.

Machine Learning Models to Classify Shiitake Mushrooms (Lentinula edodes) According to Their Geographical Origin Labeling.

The shiitake mushroom has gained popularity in the last decade, ranking second in the world for mushrooms consumed, providing consumers with a wide variety of nutritional and healthy benefits. It is often not clear the origin of these mushrooms, so it becomes of great importance to the consumers. In this research, different machine learning algorithms were developed to determine the geographical origin of shiitake mushrooms (Lentinula edodes) consumed in Korea, based on experimental data reported in the literature (δ13C, δ15N, δ18O, δ34S, and origin). Regarding the origin of shiitake in three categories (Korean, Chinese, and mushrooms from Chinese inoculated sawdust blocks), the random forest model presents the highest accuracy value (0.940) and the highest kappa value (0.908) for the validation phase. To determine the origin of shiitake mushrooms in two categories (Korean and Chinese, including mushrooms from Chinese inoculated sawdust blocks in the latter ones), the support vector machine model is chosen as the best model due to the high accuracy (0.988) and kappa (0.975) values for the validation phase. Finally, to determine the origin in two categories (Korean and Chinese, but this time including the mushrooms from Chinese inoculated sawdust blocks in the Korean ones), the best model is the random forest due to its higher accuracy value (0.952) in the validation phase (kappa value of 0.869). The accuracy values in the testing phase for the best selected models are acceptable (between 0.839 and 0.964); therefore, the predictive capacity of the models could be acceptable for their use in real applications. This allows us to affirm that machine learning algorithms would be suitable modeling instruments to determine the geographical origin of shiitake.

Physicochemical stability of bortezomib solutions for subcutaneous administration.

For the majority of cytotoxic drug preparations, such as bortezomib, the unit dose information is not available. In addition, there is a lack of information on the physicochemical stability of the pharmaceutical preparation after opening; this information is crucial for its administration to patients in successive visits, and the per-patient cost can be affected. The purpose of our proposed physicochemical stability study is to determine the shelf life of the reconstituted liquid product under refrigeration and clinical practice conditions. This evaluation was extended to both vials and ready-to-use syringes prefilled with the contents of the open vial. The stability test design includes the specified storage conditions and the critical physicochemical parameters of reconstituted injectable bortezomib. Furthermore, this approach includes the determination of impurities, the monitoring of the purity of the mean peak using a photodiode array, the control of the mass balance, the monitoring of subvisible particles using a laser diffraction analyser, and the setting of stability specifications. For the chemical stability study, the amount of bortezomib and its degradation products were determined using a stability-indicating HPLC method. The physical inspection of the samples was performed throughout the stability study, and their pH values were also monitored. Bortezomib (2.5 mg/mL) in 0.9% sodium chloride remained stable for 7 days when stored in both polypropylene syringes and vials at 5 ± 3 °C (refrigeration) and shielded from light. Additionally, it exhibits stability for 24 h under storage conditions simulating clinical use (20-30 °C and protected from light). The proposed protocol provides the stability in the vials once reconstituted and in prefilled refrigerated syringes; this protocol can be used to reduce waste and increase cost savings.