The alternative sigma factor RpoS regulates Pseudomonas aeruginosa quorum sensing response by repressing the pqsABCDE operon and activating vfr.

Pseudomonas aeruginosa is an important opportunistic pathogen. Several of its virulence-related processes, including the synthesis of pyocyanin (PYO) and biofilm formation, are controlled by quorum sensing (QS). It has been shown that the alternative sigma factor RpoS regulates QS through the reduction of lasR and rhlR transcription (encoding QS regulators). However, paradoxically, the absence of RpoS increases PYO production and biofilm development (that are RhlR dependent) by unknown mechanisms. Here, we show that RpoS represses pqsE transcription, which impacts the stability and activity of RhlR. In the absence of RpoS, rhlR transcript levels are reduced but not the RhlR protein concentration, presumably by its stabilization by PqsE, whose expression is increased. We also report that PYO synthesis and the expression of pqsE and phzA1B1C1D1E1F1G1 operon exhibit the same pattern at different RpoS concentrations, suggesting that the RpoS-dependent PYO production is due to its ability to modify PqsE concentration, which in turn modulates the activation of the phzA1 promoter by RhlR. Finally, we demonstrate that RpoS favors the expression of Vfr, which activates the transcription of lasR and rhlR. Our study contributes to the understanding of how RpoS modulates the QS response in P. aeruginosa, exerting both negative and positive regulation.

SanPy: Software for the analysis and visualization of whole-cell current-clamp recordings.

The analysis of action potentials and other membrane voltage fluctuations provides a powerful approach for interrogating the function of excitable cells. However, a major bottleneck in the interpretation of this critical data is the lack of intuitive, agreed-upon software tools for its analysis. Here, we present SanPy, an open-source and freely available software package for the analysis and exploration of whole-cell current-clamp recordings written in Python. SanPy provides a robust computational engine with an application programming interface. Using this, we have developed a cross-platform desktop application with a graphical user interface that does not require programming. SanPy is designed to extract common parameters from action potentials, including threshold time and voltage, peak, half-width, and interval statistics. In addition, several cardiac parameters are measured, including the early diastolic duration and rate. SanPy is built to be fully extensible by providing a plugin architecture for the addition of new file loaders, analysis, and visualizations. A key feature of SanPy is its focus on quality control and data exploration. In the desktop interface, all plots of the data and analysis are linked, allowing simultaneous data visualization from different dimensions with the goal of obtaining ground-truth analysis. We provide documentation for all aspects of SanPy, including several use cases and examples. To test SanPy, we performed analysis on current-clamp recordings from heart and brain cells. Taken together, SanPy is a powerful tool for whole-cell current-clamp analysis and lays the foundation for future extension by the scientific community.

CRISPR/sgRNA-directed synergistic activation mediator (SAM) as a therapeutic tool for Parkinson´s disease.

Parkinson`s disease (PD) is the second most prevalent neurodegenerative disease, and different gene therapy strategies have been used as experimental treatments. As a proof-of-concept for the treatment of PD, we used SAM, a CRISPR gene activation system, to activate the endogenous tyrosine hydroxylase gene (th) of astrocytes to produce dopamine (DA) in the striatum of 6-OHDA-lesioned rats. Potential sgRNAs within the rat th promoter region were tested, and the expression of the Th protein was determined in the C6 glial cell line. Employing pseudo-lentivirus, the SAM complex and the selected sgRNA were transferred into cultures of rat astrocytes, and gene expression and Th protein synthesis were ascertained; furthermore, DA release into the culture medium was determined by HPLC. The DA-producing astrocytes were implanted into the striatum of 6-OHDA hemiparkinsonian rats. We observed motor behavior improvement in the lesioned rats that received DA-astrocytes compared to lesioned rats receiving astrocytes that did not produce DA. Our data indicate that the SAM-induced expression of the astrocyte´s endogenous th gene can generate DA-producing astrocytes that effectively reduce the motor asymmetry induced by the lesion.

Gene replacement therapy for spinal muscular atrophy: safety and preliminary efficacy in a Brazilian cohort.

Spinal muscular atrophy (SMA) is a motor neuron disease associated with progressive muscle weakness, ventilatory failure, and reduced survival. Onasemnogene abeparvovec is the first gene replacement therapy (GT) approved to treat this condition. An observational retrospective study was conducted to assess adverse events and efficacy of GT in SMA patients. Forty-one patients with SMA (58.5% females and 80.1% SMA type 1) were included. The mean age at GT dosing was 18 (±6.4) months. Thirty-six patients (87.8%) were under previous treatment with nusinersen, and 10 (24.4%) continued nusinersen after GT. Mean CHOP-INTEND increased 13 points after 6 months and this finding did not differ between groups according to nusinersen maintenance after GT (p = 0.949). Among SMA type 1 patients, 14 (46.6%) reached the ability to sit alone. Liver transaminases elevation at least two times higher than the upper limit of normal value occurred in 29 (70.7%) patients. Thrombocytopenia occurred in 13 (31.7%) patients, and one presented thrombotic microangiopathy. Older age (>2 years) was associated with more prolonged use of corticosteroids (p = 0.021). GT is effective in SMA patients, combined nusinersen after GT did not appear to add gain in motor function and older age is associated with prolonged corticosteroid use.

Detection and evaluation of parameters influencing the identification of heterozygous-enriched regions in Holstein cattle based on SNP chip or whole-genome sequence data.

BACKGROUND: A heterozygous-enriched region (HER) is a genomic region with high variability generated by factors such as balancing selection, introgression, and admixture processes. In this study, we evaluated the genomic background of HERs and the impact of different parameters (i.e., minimum number of SNPs in a HER, maximum distance between two consecutive SNPs, minimum length of a HER, maximum number of homozygous allowed in a HER) and scenarios [i.e., different SNP panel densities and whole-genome sequence (WGS)] on the detection of HERs. We also compared HERs characterized in Holstein cattle with those identified in Angus, Jersey, and Norwegian Red cattle using WGS data. RESULTS: The parameters used for the identification of HERs significantly impact their detection. The maximum distance between two consecutive SNPs did not impact HERs detection as the same average of HERs (269.31 ± 787.00) was observed across scenarios. However, the minimum number of markers, maximum homozygous markers allowed inside a HER, and the minimum length size impacted HERs detection. For the minimum length size, the 10 Kb scenario showed the highest average number of HERs (1,364.69 ± 1,483.64). The number of HERs decreased as the minimum number of markers increased (621.31 ± 1,271.83 to 6.08 ± 21.94), and an opposite pattern was observed for the maximum homozygous markers allowed inside a HER (54.47 ± 195.51 to 494.89 ± 1,169.35). Forty-five HER islands located in 23 chromosomes with high Tajima's D values and differential among the observed and estimated heterozygosity were detected in all evaluated scenarios, indicating their ability to potentially detect regions under balancing selection. In total, 3,440 markers and 28 genes previously related to fertility (e.g., TP63, ZSCAN23, NEK5, ARHGAP44), immunity (e.g., TP63, IGC, ARHGAP44), residual feed intake (e.g., MAYO9A), stress sensitivity (e.g., SERPINA6), and milk fat percentage (e.g., NOL4) were identified. When comparing HER islands among breeds, there were substantial overlaps between Holstein with Angus (95.3%), Jersey (94.3%), and Norwegian Red cattle (97.1%), indicating conserved HER across taurine breeds. CONCLUSIONS: The detection of HERs varied according to the parameters used, but some HERs were consistently identified across all scenarios. Heterozygous genotypes observed across generations and breeds appear to be conserved in HERs. The results presented could serve as a guide for defining HERs detection parameters and further investigating their biological roles in future studies.

Modulation of neuronal activity in human centromedian nucleus during an auditory attention and working memory task.

Centromedian nucleus (CM) is one of several intralaminar nuclei of the thalamus and is thought to be involved in consciousness, arousal, and attention. CM has been suggested to play a key role in the control of attention, by regulating the flow of information to different brain regions such as the ascending reticular system, basal ganglia, and cortex. While the neurophysiology of attention in visual and auditory systems has been studied in animal models, combined single unit and LFP recordings in human have not, to our knowledge, been reported. Here, we recorded neuronal activity in the CM nucleus in 11 patients prior to insertion of deep brain stimulation electrodes for the treatment of epilepsy while subjects performed an auditory attention task. Patients were requested to attend and count the infrequent (p = 0.2) odd or "deviant" tones, ignore the frequent standard tones and report the total number of deviant tones at trial completion. Spikes were discriminated, and LFPs were band pass filtered (5-45 Hz). Average peri­stimulus time histograms and spectra were constructed by aligning on tone onsets and statistically compared. The firing rate of CM neurons showed selective, multi-phasic responses to deviant tones in 81% of the tested neurons. Local field potential analysis showed selective beta and low gamma (13-45 Hz) modulations in response to deviant tones, also in a multi-phasic pattern. The current study demonstrates that CM neurons are under top-down control and participate in the selective processing during auditory attention and working memory. These results, taken together, implicate the CM in selective auditory attention and working memory and support a role of beta and low gamma oscillatory activity in cognitive processes. It also has potential implications for DBS therapy for epilepsy and non-motor symptoms of PD, such as apathy and other disorders of attention.

Hookworm infection as a model for deepen knowledge of iron metabolism and erythropoiesis in anemia.

Over the years, there has been progress in understanding the molecular aspects of iron metabolism and erythropoiesis. However, despite research conducted both in laboratories and living organisms, there are still unanswered questions due to the complex nature of these fields. In this study we investigated the effects of hookworm infection on iron metabolism and how the hosts response to anemia is affected using hamsters infected with Ancylostoma ceylanicum as a model. Our data revealed interesting relationships between infection-induced anemia, erythropoiesis, iron metabolism, and immune modulation, such that the elevated production of erythropoietin (EPO) in renal tissue indicated intensified erythropoiesis in response to anemia. Additionally, the increased expression of the erythroferrone (ERFE) gene in the spleen suggested its involvement in iron regulation and erythropoiesis. Gene expression patterns of genes related to iron metabolism varied in different tissues, indicating tissue-specific adaptations to hypoxia. The modulation of pro-inflammatory and anti-inflammatory cytokines highlighted the delicate balance between immune response and erythropoiesis. Data derived from the investigation of changes induced in iron metabolism and stress erythropoiesis following anemia aid in our understanding of mechanisms related to blood spoliation and anemia, which could potentially be extrapolated or compared to other types or causes of anemia. These findings also contribute to our understanding of the pathophysiology of erythropoiesis in the context of blood loss.

Existing and Emerging Technologies for Therapeutic Monitoring of Lithium: A Scoping Review.

BACKGROUND/PURPOSE: Lithium is an effective psychoactive drug. It has a narrow therapeutic margin, with subtherapeutic levels or intoxication commonly occurring. Therapeutic drug monitoring (TDM) of lithium has several barriers. This scoping review aims to describe and analyze existing and emerging technologies for lithium TDM and to describe the lithium quantification parameters (precision, accuracy, detection limit) attributed to each technology. METHOD: PubMed, Scopus, Web of Science, and Google Scholar were searched. Studies that described lithium quantification and complied with PRISMA-ScR guidelines were included. Articles selection was conducted by 2 researchers. Good precision was defined if its relative standard deviation <3%; acceptable, from 3% to 5%; and low, >5%. Accuracy was considered good if the error <5%; acceptable, 5%1 to 0%; and low if it was >10%. RESULTS: Of the 2008 articles found, 22 met the inclusion criteria. Of these, 14 studies concerned laboratory devices, in which precision was found to be low in one third of cases, and half had good precision. Accuracy of one third was good, another third was low, and the remaining third did not report accuracy. The other 8 studies concerned portable devices, in which precision was low in more than 60% of the cases and good in 25% of the studies. Accuracy was low in 50% of the cases, and good in just over a third. Limits of detection included the therapeutic range of lithium in all studies. CONCLUSIONS: Among emerging technologies for lithium TDM, precision and accuracy remain a challenge, particularly for portable devices.

H2OAthletes study protocol: effects of hydration changes on neuromuscular function in athletes.

We aim to understand the effects of hydration changes on athletes' neuromuscular performance, on body water compartments, fat-free mass hydration and hydration biomarkers and to test the effects of the intervention on the response of acute dehydration in the hydration indexes. The H2OAthletes study (clinicaltrials.gov ID: NCT05380089) is a randomised controlled trial in thirty-eight national/international athletes of both sexes with low total water intake (WI) (i.e. < 35·0 ml/kg/d). In the intervention, participants will be randomly assigned to the control (CG, n 19) or experimental group (EG, n 19). During the 4-day intervention, WI will be maintained in the CG and increased in the EG (i.e. > 45·0 ml/kg/d). Exercise-induced dehydration protocols with thermal stress will be performed before and after the intervention. Neuromuscular performance (knee extension/flexion with electromyography and handgrip), hydration indexes (serum, urine and saliva osmolality), body water compartments and water flux (dilution techniques, body composition (four-compartment model) and biochemical parameters (vasopressin and Na) will be evaluated. This trial will provide novel evidence about the effects of hydration changes on neuromuscular function and hydration status in athletes with low WI, providing useful information for athletes and sports-related professionals aiming to improve athletic performance.

Pentavalent and poliomyelitis vaccines: a review of the vaccination coverage in Brazilian children in the last 10 years.

The pentavalent and poliomyelitis vaccines are provided to children through the National Immunization Program in Brazil. For the last few years, this program has been responsible for eradication of various diseases in the country. To describe the vaccination coverage of the pentavalent vaccine and the polio vaccine in Brazil from 2013 to 2022. This is a retrospective and descriptive study with a time series component in which we analyzed vaccination coverage rates of pentavalent and poliomyelitis in the national vaccination program of Brazil from 2013 to 2022. The collected data were arranged in spreadsheets, analyzed, and presented graphically. In general, the coverage of the poliomyelitis and pentavalent vaccines has declined in all regions of Brazil, with significant differences each year. Although there has been a decrease in vaccination coverage in all regions and states, the most affected regions are the North and Northeast. There has been a significant decrease in polio and pentavalent vaccine coverage in recent years, and the rate of decrease differs in different parts of the country.

Genotyping of Toxoplasma gondii in domestic animals from Campeche, México, reveals virulent genotypes and a recombinant ROP5 allele.

Toxoplasma gondii has at least 318 genotypes distributed worldwide, and tropical regions usually have greater genetic diversity. Campeche is a state located in the southeastern region of México and has favourable climate conditions for the replication and dissemination of this protozoan, similar to those in South American countries where broad genetic diversity has been described. Thus, in this study, 4 T. gondii isolates were obtained from tissues of stray dogs and free-range chickens in Campeche, México, and were genotyped by Mn-PCR-RFLP with 10 typing markers (SAG1, altSAG2, SAG3, BTUB, GRA6, c22-8, c29-2, L358, PK1 and Apico) and 5 virulence markers (CS3, ROP16, ROP17, ROP18 and ROP5) to provide new information about the distribution and virulence prediction of T. gondii genotypes. Two isolates of T. gondii genotype #116 and 2 of genotype #38 were obtained from stray dogs and chickens, respectively. The parasite load found in these species was between <50 and more than 35 000 tachyzoites per mg of tissue. Virulence marker genotyping revealed a recombinant 1&3 ROP5 RFLP pattern in 2 ToxoDB #116 isolates with no prediction of virulence in a murine model, while in the 2 ToxoDB #38 isolates, the ROP18/ROP5 combination predicted high virulence. Considering all the typed markers, there is a predominance of type I and III alleles, as constantly reported for the isolates characterized in various regions of México. It is crucial to determine their phenotype to corroborate the genetic virulence profile of the T. gondii isolates obtained in this study.

In situ interaction between the hormone 17α-ethynylestradiol and the liquid-ordered phase composed of the lipid rafts sphingomyelin and cholesterol.

Hormone treatments are frequently associated with cardiovascular diseases and cancers in women. Additionally, the detrimental effects of their presence as contaminants in water remain a concern. The transport of hormones through cell membranes is essential for their biological action, but investigating cell permeability is challenging owing to the experimental difficulty in dealing with whole cells. In this paper, we study the interaction of the synthetic hormone 17α-ethynylestradiol (EE2) with membrane models containing the key raft components sphingomyelin (SM) and cholesterol (Chol). The models consisted of Langmuir monolayers and giant unilamellar vesicles (GUVs) that represent bilayers. EE2 induced expansion of SM monolayers upon interacting with the non-hydrated amide group of SM head, but it had practically no effect on SM GUVs because these group are not available for interaction in bilayers. In contrast, EE2 interacted with hydrated phosphate group (PO2-) and amide group of SM/Chol mixture monolayer, which could explain the loss in phase contrast of liquid-ordered GUVs suggesting pore formation. A comparison with reported EE2 effects on GUVs in the fluid phase, for which no loss in phase contrast was observed, indicates that the liquid-ordered phase consisting of lipid rafts is relevant to be associated with the changes on cell permeability caused by the hormones.

Left knee septic monoarthritis in a pediatric patient due to shewanella putrefaciens: case report and literature review.

BACKGROUND: Shewanella putrefaciens is a gram-negative, nonfermenting, oxidase-positive, hydrogen sulfide-producing bacillus and a halophilic bacterium, known for causing unusual infections in humans and often regarded as an opportunistic pathogen. Its diverse symptoms have a significant impact on human health, with 260 documented disorders reported in the literature over the last 40 years, highlighting its potential danger. CASE PRESENTATION: We present the case of a previously healthy 15-year-old male patient who sustained a self-inflicted sharp-object injury while working in the field, resulting in secondary septic monoarthritis due to Shewanella putrefaciens. CONCLUSIONS: This case highlights the bacteriological and clinical characteristics, as well as the antibiogram, of Shewanella spp. Given the recent increase in notifications of Shewanella infections, predominantly by S. algae and S. putrefaciens, it is essential to consider these pathogens in patients with a history of contact with bodies of water. Special attention must be paid to their resistance patterns in patient management to prevent the development of intrinsic antimicrobial resistance.

Spanish validation of the short version of the racing and crowded thoughts questionnaire (RCTQ-13).

BACKGROUND: The Racing and Crowded Thoughts Questionnaire (RCTQ-13) is the most widely used specific scale for the measurement of racing thoughts, but there is currently no Spanish version that allow the evaluation in Spanish-speaking patients. The objective of this study is to translate, adapt, and validate the RCTQ-13 in a Colombian population with affective disorders. METHODS: The questionnaire was translated and back-translated, and corrections were implemented following a pilot test to improve comprehensibility. We included patients with Bipolar I Disorder and with Major depressive disorder seen in three centers in the city of Medellín, Colombia. We evaluate structural validity with confirmatory factor analysis, internal consistency, and test-retest reliability. Construct validity was also assessed with the comparison between euthymic, maniac, and depressive episodes and the correlation with worry, rumination, and mania scales. Responsiveness was measured 1 month after the first evaluation. Based on item response theory (IRT), we also estimated item difficulty, discrimination, and fit using a generalized partial credit model. RESULTS: Two hundred fifty subjects were included. Confirmatory factor analysis revealed that the three-factor structure of the scale was appropriate. Internal consistency was adequate for the entire scale (Cronbach's alpha = 0.95, 95% CI: 0.94-0.96) and for each factor. Test-retest reliability was good (intraclass correlation coefficient = 0.82, 95%IC: 0.70-0.88). For construct validity, we observed differences between patients with different types of affective episodes, a moderate positive correlation with the Penn State Worry Scale (r = 0.55) and the Ruminative Response Scale (r = 0.42), and a low negative correlation with the Young Mania Rating Scale (r = - 0.10). Responsiveness was proved to be adequate. Under IRT, the response thresholds for the response options are organized for all items. The infit was adequate for all items and the outfit was acceptable. CONCLUSIONS: The Spanish version of the RCTQ-13 is a reliable, valid, and responsive scale and can be used for the clinical assessment of the construct of racing and crowded thoughts in patients with the spectrum of affective disorders in whom this experience can be expressed with different nuances. Further research is needed to expand the relationship with rumination and worry.

The conundrum of differentiating Cushing's syndrome from non-neoplastic hypercortisolism: a systematic review and meta-analysis.

CONTEXT: Once hypercortisolemia is confirmed, differential diagnosis between Cushing's syndrome (CS) due to neoplastic endogenous hypercortisolism and non-neoplastic hypercortisolism (NNH, pseudo-Cushing's syndrome) is crucial. Due to worldwide corticotropin-releasing hormone (CRH) unavailability, accuracy of alternative tests to dexamethasone (Dex)-CRH, is clearly needed. OBJECTIVE: Assess the diagnostic accuracy of Dex-CRH test, desmopressin stimulation test, midnight serum cortisol (MSC), and late-night salivary cortisol (LNSC) levels to distinguish CS from NNH. METHODS: Articles through March 2022 were identified from Scopus, Web of Science, MEDLINE, EMBASE, and PubMed. All steps through the systematic review were performed independently and in duplicate and strictly adhered to the updated PRISMA-DTA checklist. DATA SYNTHESIS: A total of 24 articles (1900 patients) were included. Dex-CRH had a pooled sensitivity and specificity of 91% (95%CI 87-94%; I2 0%) and 82% (73-88%; I2 50%), desmopressin test 86% (81-90%; I2 28%) and 90% (84-94%; I2 15%), MSC 91% (85-94%; I2 66%) and 81% (70-89%; I2 71%), and LNSC 80% (67-89%; I2 57%) and 90% (84-93%; I2 21%), respectively. Summary receiver operating characteristics areas under the curve were Dex-CRH 0.949, desmopressin test 0.936, MSC 0.942, and LNSC 0.950 without visual or statistical significance. The overall risk of studies bias was moderate. CONCLUSION: Dex-CRH, the desmopressin stimulation test, and MSC have similar diagnostic accuracy, with Dex-CRH and MSC having slightly higher sensitivity, and the desmopressin test being more specific. LNSC was the least accurate, probably due to high heterogeneity, intrinsic variability, different assays, and lack of consistent reported cutoffs. When facing this challenging differential diagnosis, the results presented here should increase clinicians' confidence when deciding which test to perform.

Climacteric and fibromyalgia: a review.

Fibromyalgia (FM) and climacteric conditions share common epidemiological and clinical features, with FM symptoms often beginning during menopause. Musculoskeletal pain, arthralgia, myalgia and other symptoms are frequently seen in both conditions. Some research suggests a link between the cessation of sex hormones and FM symptoms. Women with FM tend to experience more severe symptoms after menopause, and the severity of FM symptoms can worsen in women who have had a hysterectomy with or without oophorectomy. Despite these similarities, it is essential to treat FM and climacteric conditions separately and follow established guidelines for management. However, it is also important to recognize that both conditions can coexist in the same patient. It is crucial to note that there is limited evidence supporting the effectiveness of menopausal hormone therapy for primary FM management. Therefore, menopausal hormone therapy should not be recommended for FM unless the patient also has climacteric syndrome.

Exposure of cinnamyl alcohol in co-culture of BEAS-2B and dendritic cells: Interaction between CYP450 and cytokines.

The prevalence of fragrances in various hygiene products contributes to their sensorial allure. However, fragrances can induce sensitization in the skin or respiratory system, and the mechanisms involved in this process are incompletely understood. This study investigated the intricate mechanisms underlying the fragrance's effects on sensitization response, focusing on the interplay between CYP450 enzymes, a class of drug-metabolizing enzymes, and the adaptive immune system. Specifically, we assessed the expression of CYP450 enzymes and cytokine profiles in culture of BEAS-2B and mature dendritic cells (mDC) alone or in co-culture stimulated with 2 mM of a common fragrance, cinnamyl alcohol (CA) for 20 h. CYP1A1, CYP1A2, CYP1B1, CYP2A6, and CYP2A13 were analyzed by RT-PCR and IL-10, IL-12p70, IL-18, IL-33, and thymic stromal lymphopoietin (TSLP) by Cytometric Bead Array (CBA). Through RT-PCR analysis, we observed that CA increased CYP1A2 and CYP1B1 expression in BEAS-2B, with a further increased in BEAS-2B-mDC co-culture. Additionally, exposure to CA increased IL-12p70 levels in mDC rather than in BEAS-2B-mDC co-culture. In regards to IL-18, level was higher in BEAS-2B than in BEAS-2B-mDC co-culture. A positive correlation between the levels of IL-10 and CYP1B1 was found in mDC-CA-exposed and between IL-12p70 and CYP1A1 was found in BEAS-2B after CA exposure. However, IL-12p70 and CYP1A2 as well as IL-18, IL-33, and CYP1A1 levels were negative, correlated mainly in co-culture control. These correlations highlight potential immunomodulatory interactions and complex regulatory relationships. Overall, exposure to CA enhances CYP450 expression, suggesting that CA can influence immune responses by degrading ligands on xenosensitive transcription factors.

Crystal structure of dihydroneopterin aldolase from Mycobacterium tuberculosis associated with 8-mercaptoguanine, and development of novel S8-functionalized analogues as inhibitors: Synthesis, enzyme inhibition, in vitro toxicity and antitubercular activity.

The crystallographic structure of the FolB enzyme from Mycobacterium tuberculosis (MtFolB), complexed with its inhibitor 8-mercaptoguanine (8-MG), was elucidated at a resolution of 1.95 Å. A novel series of S8-functionalized 8-MG derivatives were synthesised and evaluated as in vitro inhibitors of dihydroneopterin aldolase (DHNA, EC 4.1.2.25) activity of MtFolB. These compounds exhibited IC50 values in the submicromolar range. Evaluation of the activity for five compounds indicated their inhibition mode and inhibition constants. Molecular docking analyses were performed to determine the enzyme-inhibitor intermolecular interactions and ligand conformations upon complex formation. The inhibitory activities of all compounds against the M. tuberculosis H37Rv strain were evaluated. Compound 3e exhibited a minimum inhibitory concentration in the micromolar range. Finally, Compound 3e showed no apparent toxicity in both HepG2 and Vero cells. The findings presented herein will advance the quest for novel, specific inhibitors targeting MtFolB, an attractive molecular target for TB drug development.

Characteristics and onset of presentation of pediatric stiff skin syndrome: A retrospective cohort study of 11 patients in a tertiary care center.

BACKGROUND/OBJECTIVE: Stiff skin syndrome (SSS) is a rare disorder characterized by "rock hard" indurated skin affecting different body parts. The localized variant poses a diagnostic challenge, as it is frequently mistaken for other inflammatory connective tissue disorders. The aim of this study is to provide insightful clinical, radiologic and diagnostic data that might prove useful for the evaluation, management and treatment of pediatric patients with segmental SS. METHODS: This single-center cohort study included patients ≤18 years diagnosed with localized SSS from 1988 to 2021 in a quaternary pediatric healthcare center in Toronto, Canada. Data included demographics, clinical, histopathologic and radiologic features, treatments, and clinical course. Data were summarized with descriptive statistics (mean, standard deviation, medians, interquartile ranges [IQRs]) and frequencies. RESULTS: A total of 11 patients were included. The sclerotic changes were measured clinically and radiologically, by a total of 16 imaging studies: 13 magnetic resonance imaging (MRI) and 3 ultrasound. MRI readings showed abnormal high signal intensity of the affected tissue correlating with the anatomical site of involvement in all cases, specifically, in the shoulder/pelvic girdle with limb extension. Shear wave ultrasound elastography (SWE) demonstrated higher values within the dermis compared to the control site. CONCLUSION: The presence of segmental sclerotic changes that affects the pelvic/shoulder girdle with extension to the extremities, in the absence of inflammation on biopsy and abnormal signaling intensity on imaging is suggestive of SSS. Skin SWE is a feasible, noninvasive, and objective instrument to evaluate and monitor sclerotic changes overtime, it could be potentially extrapolated to other pediatric skin sclerotic conditions.

Single dominant lesion in capillary malformation-arteriovenous malformation (CM-AVM) RASA1 syndrome.

We report two cases with localized vascular malformations clinically resembling the "dominant lesion" seen in capillary malformation-arteriovenous malformation (CM-AVM) syndrome, however, lacking germline RASA1 variants but presenting double somatic RASA1 variants in affected tissue. Both patients presented with localized and superficial high-flow vascular malformations were treated with surgery and laser therapy and showed partial resolution. The study underscores the rarity of somatic RASA1 variants, contributes to understanding the "second-hit" pathophysiology in vascular lesions, and emphasizes the significance of clinical distinctions and genotyping for accurate diagnoses, offering implications for diagnosis, prognosis, and genetic counseling.