Confined Water Dynamics in the Scaffolds of Polylactic Acid.

Resorbable polylactic acid (PLA) ultrathin fibers have been applied as scaffolds for tissue engineering applications due to their micro- and nanoporous structure that favor cell adhesion, besides inducing cell proliferation and upregulating gene expression related to tissue regeneration. Incorporation of multiwalled carbon nanotubes into PLA fibers has been reported to increase the mechanical properties of the scaffold, making them even more suitable for tissue engineering applications. Ideally, scaffolds should be degraded simultaneously with tissue growth. Hydration and swelling are factors related to scaffold degradation. Hydration would negatively impact the mechanical properties since PLA shows hydrolytic degradation. Water absorption critically affects the catalysis and allowance of the hydrolysis reactions. Moreover, either mass transport and chemical reactions are influenced by confined water, which is an unexplored subject for PLA micro- and nanoporous fibers. Here, we probe and investigate confined water onto highly porous PLA microfibers containing few amounts of incorporated carbon nanotubes by Fourier transform infrared (FTIR) spectroscopy. A hydrostatic pressure was applied to the fibers to enhance the intermolecular interactions between water molecules and C=O groups from polyester bonds, which were evaluated over the wavenumber between 1600 and 2000 cm-1. The analysis of temperature dependence of FTIR spectra indicated the presence of confined water which is characterized by a non-Arrhenius to Arrhenius crossover at T0 = 190 K for 1716 and 1817 cm-1 carbonyl bands of PLA. These bands are sensitive to a hydrogen bond network of confined water. The relevance of our finding relies on the challenge detecting confined water in hydrophobic cavities as in the PLA one. To the best of our knowledge, we present the first report referring the presence of confined water in a hydrophobic scaffold as PLA for tissue engineering. Our findings can provide new opportunities to understand the role of confined water in tissue engineering applications. For instance, we argue that PLA degradation may be affected the most by confined water. PLA degradation involves hydrolytic and enzymatic degradation reactions, which can both be sensitive to changes in water properties.

Amino acid supplementation confers protection to red blood cells before Plasmodium falciparum bystander stress.

ABSTRACT: Malaria is a highly oxidative parasitic disease in which anemia is the most common clinical symptom. A major contributor to the malarial anemia pathogenesis is the destruction of bystander, uninfected red blood cells (RBCs). Metabolic fluctuations are known to occur in the plasma of individuals with acute malaria, emphasizing the role of metabolic changes in disease progression and severity. Here, we report conditioned medium from Plasmodium falciparum culture induces oxidative stress in uninfected, catalase-depleted RBCs. As cell-permeable precursors to glutathione, we demonstrate the benefit of pre-exposure to exogenous glutamine, cysteine, and glycine amino acids for RBCs. Importantly, this pretreatment intrinsically prepares RBCs to mitigate oxidative stress.

Surgical treatment of canine urethral prolapse via urethropexy or resection and anastomosis.

OBJECTIVES: The objective was to report and compare the complications and recurrence rates of urethral prolapse in dogs when treated with urethropexy, resection and anastomosis or a combined surgical technique. STUDY DESIGN: Retrospective study. MATERIALS AND METHODS: A total of 86 dogs were identified from the medical records of 10 veterinary referral hospitals from February 2012 and October 2022. Dogs were included if they underwent surgery for a urethral prolapse at first presentation. Complications were classified as minor or major based on the necessity of further surgical intervention. Complications leading to death were also considered major complications. RESULTS: Seventy-nine dogs were included, urethropexy (n=44), resection and anastomosis (n=27) and a combined surgical technique (n=8). Minor complications were identified in 41 of 79 dogs (51.9%): urethropexy 19 of 44 (43.2%), resection and anastomosis 18 of 27 (66.6%) and a combined surgical technique four of eight (50%). Major complications occurred in 23 dogs (29.1%), of which 21 were recurrence (26.6%). Recurrence occurred in 17 of 44 dogs following a urethropexy (38.6%), three of 27 dogs following resection and anastomosis (11.1%) and one of eight dogs treated with a combined surgical technique (12.5%). Recurrence of a urethral prolapse was significantly more likely following urethropexy in comparison to resection and anastomosis. CLINICAL SIGNIFICANCE: Resection and anastomosis was associated with a lower recurrence rate in comparison to urethropexy for the surgical treatment of urethral prolapse. Based on these results, we concluded that resection and anastomosis may be preferable to urethropexy for treatment of urethral prolapse at first presentation. Urethropexy, and resection and anastomosis combined surgical technique was associated with low recurrence rate; however, further studies will be needed to clarify if it provides any benefit over resection and anastomosis.

Compatibility Assessment of Novel Orodispersible Film Vehicle for Personalized Medicine with Selected Active Pharmaceutical Ingredients.

Orodispersible films (ODFs) are solid pharmaceutical forms for rapid local or systemic release of active ingredients. They are formed by a water-soluble polymer film that hydrates rapidly, adhering and dissolving immediately when placed on the tongue or in the oral cavity. In this paper, we describe the compatibility and disintegration times of compounded ODFs using OrPhylloTM, a new ready-to-use-vehicle, and APIs from different pharmacological classes, such as 5-hydroxytryptophan (5-HTP) 50 mg, bromopride 5 mg, coenzyme Q10 20 mg, melatonin 3 mg, resveratrol 5 mg, tadalafil 10 mg, vitamin B12 1 mg, or vitamin D3 2000 UI. ODFs were compounded and, subsequently, the samples were assayed using HPLC at initial (t = 0), 7 days (t = 7), 14 days (t = 14), 30 days (t = 30), 60 days (t = 60), 90 days (t = 90), 120 days (t = 120), 150 days (t = 150), and 180 days (t = 180) after compounding. Given the percentage of recovery of the APIs within the films, the beyond-use date of the final products (API + vehicle) was at least 90 days for vitamin D3, 150 days for bromopride and 5-HTP, and 180 days for coenzyme Q10, tadalafil, vitamin B12, resveratrol, and melatonin, when stored at room temperature. The average disintegration time was 46.22 s. This suggests that the OrPhylloTM vehicle is suitable for compounding ODFs with APIs from different pharmacological classes, with good compatibility and fast disintegration.

Goldfish adiponectin: (I) molecular cloning, tissue distribution, recombinant protein expression, and novel function as a satiety factor in fish model.

Adiponectin (AdipoQ) is an adipokine involved in glucose homeostasis and lipid metabolism. In mammals, its role in appetite control is highly controversial. To shed light on the comparative aspects of AdipoQ in lower vertebrates, goldfish was used as a model to study feeding regulation by AdipoQ in fish species. As a first step, goldfish AdipoQ was cloned and found to be ubiquitously expressed at the tissue level. Using sequence alignment, protein modeling, phylogenetic analysis and comparative synteny, goldfish AdipoQ was shown to be evolutionarily related to its fish counterparts and structurally comparable with AdipoQ in higher vertebrates. In our study, recombinant goldfish AdipoQ was expressed in E. coli, purified by IMAC, and confirmed to be bioactive via activation of AdipoQ receptors expressed in HepG2 cells. Feeding in goldfish revealed that plasma levels of AdipoQ and its transcript expression in the liver and brain areas involved in appetite control including the telencephalon, optic tectum, and hypothalamus could be elevated by food intake. In parallel studies, IP and ICV injection of recombinant goldfish AdipoQ in goldfish was effective in reducing foraging behaviors and food consumption. Meanwhile, transcript expression of orexigenic factors (NPY, AgRP, orexin, and apelin) was suppressed with parallel rises in anorexigenic factors (POMC, CART, CCK, and MCH) in the telencephalon, optic tectum and/or hypothalamus. In these brain areas, transcript signals for leptin receptor were upregulated with concurrent drops in the NPY receptor and ghrelin receptors. In the experiment with IP injection of AdipoQ, transcript expression of leptin was also elevated with a parallel drop in ghrelin mRNA in the liver. These findings suggest that AdipoQ can act as a novel satiety factor in goldfish. In this case, AdipoQ signals (both central and peripheral) can be induced by feeding and act within the brain to inhibit feeding behaviors and food intake via differential regulation of orexigenic/anorexigenic factors and their receptors. The feeding inhibition observed may also involve the hepatic action of AdipoQ by modulation of feeding regulators expressed in the liver.

A New Method for Estimating Glomerular Filtration Rate in Rhesus Macaques (Macaca mulatta).

Late effects of total- or partial-body irradiation include chronic kidney injury (CKI), which increases morbidity and mortality. Glomerular filtration rate (GFR) is the gold standard measure of kidney function. Renal function markers, such as blood urea nitrogen (BUN) and serum creatinine (Cr), may not be higher than reference ranges until 50% or more of nephrons are affected. Currently available methods to measure GFR are difficult and expensive, requiring multiple blood draws or timed urine collections, but their use can provide a framework for the development of simpler GFR estimates. The measurement of iohexol clearance is a validated tool used to determine GFR in veterinary patients. In this study, we aimed to determine if the Schwartz formula as used in human pediatric medicine can estimate GFR in rhesus macaques. We hypothesized that iohexol-GFR would correlate with the Schwartz formula-estimated GFR (eGFR) in irradiated and non-irradiated rhesus macaques. Twelve rhesus macaques [age 5-14 years (mean 7 years); 5 females, 7 males] with a range of BUN levels were selected for comparison to 4 non-irradiated controls (2 females, 2 males). Irradiated animals were divided by BUN into 3 groups: BUN ≤20 mg/dL (n = 4), BUN >20-24 mg/dL (n = 4), and BUN ≥25 mg/dL (n = 4). Baseline serum chemistry and urinalysis were used to assess renal function. For measurement of GFR, macaques were maintained under general anesthesia and received an intravenous injection of iohexol (2 mL/kg, 300 mg I/mL). Whole blood was collected at 10, 30, 60 and 90 min post-iohexol injection. Plasma iohexol concentrations were determined by mass spectrometry. GFR was calculated from the peak iohexol concentration and trapezoidal area under the curve (tAUC). The iohexol-GFR significantly correlated with the Schwartz formula-eGFR. In macaques with renal irradiation doses below 6 Gy, GFR was higher for males than females. GFR was lower in macaques with renal irradiation doses greater than 6 Gy compared to macaques with renal doses less than 6 Gy. We conclude that use of the Schwartz formula can provide a rapid, non-invasive, cost-effective, and accurate estimation of GFR to aid in the clinical assessment of renal function in irradiated rhesus macaques.

Differential regulation of hepatic SH3 domain binding kinase 1 (SBK1) expression in mouse and goldfish.

SH3 domain binding kinase 1 (SBK1) is a serine/threonine kinase that belongs to the new kinase family (NFK) with limited information on its function. Previous studies reported that SBK1 plays a role in memory formation, lipid metabolism, and cancer cell progression. Nevertheless, the regulatory mechanism of Sbk1 expression in various tissues remains unknown. We report here that Sbk1 expression in mouse hepatocytes was downregulated by glucocorticoid, whereas saturated and unsaturated fatty acids were stimulators of Sbk1 expression. The regulatory role of glucocorticoid and fatty acid was further confirmed by the Sbk1 promoter assay, which aligned with the presence of several glucocorticoid-response elements (GRE) and peroxisome proliferator responsive elements (PPRE) in the mouse Sbk1 promoter. The inhibitory effect of glucocorticoids on hepatic Sbk1 expression and protein content could also be demonstrated in vivo after prednisolone injection. Moreover, the expression of SBK1 in goldfish (gfSBK1) was also sensitive to glucocorticoid suppression as their mouse orthologues. In contrast, insulin had a differential action on SBK1 expression that it promoted the expression of all SBK1 isoforms in the goldfish hepatocytes but inhibited Sbk1 expression in the mouse hepatocytes. Together, our findings indicate that SBK1 expression is hormone- and nutrient-sensitive with a species-specific response.

ABHD4 regulates adipocyte differentiation in vitro but does not affect adipose tissue lipid metabolism in mice.

Alpha/beta hydrolase domain-containing protein 4 (ABHD4) catalyzes the deacylation of N-acyl phosphatidyl-ethanolamine (NAPE) and lyso-NAPE to produce glycerophospho-N-acyl ethanolamine (GP-NAE). Through a variety of metabolic enzymes, NAPE, lyso-NAPE, and GP-NAE are ultimately converted into NAE, a group of bioactive lipids that control many physiological processes including inflammation, cognition, food intake, and lipolysis (i.e., oleoylethanolamide or OEA). In a diet-induced obese mouse model, adipose tissue Abhd4 gene expression positively correlated with adiposity. However, it is unknown whether Abhd4 is a causal or a reactive gene to obesity. To fill this knowledge gap, we generated an Abhd4 knockout (KO) 3T3-L1 pre-adipocyte. During adipogenic stimulation, Abhd4 KO pre-adipocytes had increased adipogenesis and lipid accumulation, suggesting Abhd4 is responding to (a reactive gene), not contributing to (not a causal gene), adiposity, and may serve as a mechanism for protecting against obesity. However, we did not observe any differences in adiposity and metabolic outcomes between whole-body Abhd4 KO or adipocyte-specific Abhd4 KO mice and their littermate control mice (both male and female) on chow or a high-fat diet. This might be because we found that deletion of Abhd4 did not affect NAE such as OEA production, even though Abhd4 was highly expressed in adipose tissue and correlated with fasting adipose OEA levels and lipolysis. These data suggest that ABHD4 regulates adipocyte differentiation in vitro but does not affect adipose tissue lipid metabolism in mice despite nutrient overload, possibly due to compensation from other NAPE and NAE metabolic enzymes.

Amino acid supplementation confers protection to red blood cells prior to Plasmodium falciparum bystander stress.

Malaria is a highly oxidative parasitic disease in which anemia is the most common clinical symptom. A major contributor to malarial anemia pathogenesis is the destruction of bystander, uninfected red blood cells. Metabolic fluctuations are known to occur in the plasma of individuals with acute malaria, emphasizing the role of metabolic changes in disease progression and severity. Here, we report that conditioned media from Plasmodium falciparum culture induces oxidative stress in healthy uninfected RBCs. Additionally, we show the benefit of amino acid pre-exposure for RBCs and how this pre-treatment intrinsically prepares RBCs to mitigate oxidative stress. Key points: Intracellular ROS is acquired in red blood cells incubated with Plasmodium falciparum conditioned media Glutamine, cysteine, and glycine amino acid supplementation increased glutathione biosynthesis and reduced ROS levels in stressed RBCs.

High Genetic Diversity of Carbapenem-Resistant Acinetobacter baumannii Isolates Recovered in Nigerian Hospitals in 2016 to 2020.

Acinetobacter baumannii causes difficult-to-treat infections mostly among immunocompromised patients. Clinically relevant A. baumannii lineages and their carbapenem resistance mechanisms are sparsely described in Nigeria. This study aimed to characterize the diversity and genetic mechanisms of carbapenem resistance among A. baumannii strains isolated from hospitals in southwestern Nigeria. We sequenced the genomes of all A. baumannii isolates submitted to Nigeria's antimicrobial resistance surveillance reference laboratory between 2016 and 2020 on an Illumina platform and performed in silico genomic characterization. Selected strains were sequenced using the Oxford Nanopore technology to characterize the genetic context of carbapenem resistance genes. The 86 A. baumannii isolates were phylogenetically diverse and belonged to 35 distinct Oxford sequence types (oxfSTs), 16 of which were novel, and 28 Institut Pasteur STs (pasSTs). Thirty-eight (44.2%) isolates belonged to none of the known international clones (ICs). Over 50% of the isolates were phenotypically resistant to 10 of 12 tested antimicrobials. The majority (n = 54) of the isolates were carbapenem resistant, particularly the IC7 (pasST25; 100%) and IC9 (pasST85; >91.7%) strains. blaOXA-23 (34.9%) and blaNDM-1 (27.9%) were the most common carbapenem resistance genes detected. All blaOXA-23 genes were carried on Tn2006 or Tn2006-like transposons. Our findings suggest that a 10-kb Tn125 composite transposon is the primary means of blaNDM-1 dissemination. Our findings highlight an increase in blaNDM-1 prevalence and the widespread transposon-facilitated dissemination of carbapenemase genes in diverse A. baumannii lineages in southwestern Nigeria. We make the case for improving surveillance of these pathogens in Nigeria and other understudied settings. IMPORTANCE Acinetobacter baumannii bacteria are increasingly clinically relevant due to their propensity to harbor genes conferring resistance to multiple antimicrobials, as well as their ability to persist and disseminate in hospital environments and cause difficult-to-treat nosocomial infections. Little is known about the molecular epidemiology and antimicrobial resistance profiles of these organisms in Nigeria, largely due to limited capacity for their isolation, identification, and antimicrobial susceptibility testing. Our study characterized the diversity and antimicrobial resistance profiles of clinical A. baumannii in southwestern Nigeria using whole-genome sequencing. We also identified the key genetic elements facilitating the dissemination of carbapenem resistance genes within this species. This study provides key insights into the clinical burden and population dynamics of A. baumannii in hospitals in Nigeria and highlights the importance of routine whole-genome sequencing-based surveillance of this and other previously understudied pathogens in Nigeria and other similar settings.

Hydrogel Based on Nanoclay and Gelatin Methacrylate Polymeric Matrix as a Potential Osteogenic Application.

A nanocomposite hydrogel has potentially applicability in the induction of osteogenesis. The hydrogel was synthesized using 1% gelatin methacrylate (GelMA), a biodegradable and bioactive polymer containing the structure of gelatin, denatured collagen derived from the extracellular bone matrix, and 6% laponite (Lap), a synthetic phyllosilicate of nanosized particles. Initially, 0.6 g of Lap was added to deionized water, and then a solution of GelMA/Igarcure was added under stirring and UV light for crosslinking. The spectra in the Fourier-transform infrared region showed bands that indicate the interaction between gelatin and methacrylate anhydride. X-ray diffraction patterns confirmed the presence of Lap and GelMA in the hydrogel. The thermogravimetric analysis suggested an increase in the thermal stability of the hydrogel with the presence of clay mineral. Rheological analysis showed that the hydrogel had a viscosity that allowed its injectability. The hydrogel did not show acute toxicity at any of the concentrations tested according to the Artemia salina lethality test. It showed cell viability more significant than 80% in the MTT test, which makes it suitable for in vivo osteogenic induction tests. The cell differentiation test showed the differentiation of stem cells into osteogenic cells. It indicates a material with the potential for osteogenic induction and possible application in bone tissue engineering.

A Chromosome-level assembly of the Japanese eel genome, insights into gene duplication and chromosomal reorganization.

Japanese eels (Anguilla japonica) are commercially important species, harvested extensively for food. Currently, this and related species (American and European eels) are challenging to breed on a commercial basis. As a result, the wild stock is used for aquaculture. Moreover, climate change, habitat loss, water pollution, and altered ocean currents affect eel populations negatively. Accordingly, the International Union for Conservation of Nature lists Japanese eels as endangered and on its red list. Here we presented a high-quality genome assembly for Japanese eels and demonstrated that large chromosome reorganizations occurred in the events of third-round whole-genome duplications (3R-WRDs). Several chromosomal fusions and fissions have reduced the ancestral protochromosomal number of 25 to 19 in the Anguilla lineage. A phylogenetic analysis of the expanded gene families showed that the olfactory receptors (group δ and ζ genes) and voltage-gated Ca2+ channels expanded significantly. Both gene families are crucial for olfaction and neurophysiology. Additional tandem and proximal duplications occurred following 3R-WGD to acquire immune-related genes for an adaptive advantage against various pathogens. The Japanese eel assembly presented here can be used to study other Anguilla species relating to evolution and conservation.

Nanohydroxyapatite/Titanate Nanotube Composites for Bone Tissue Regeneration.

Strategies for the production of new nanocomposites that promote bone tissue regeneration are important, particularly those that enhance the osteoinduction of hydroxyapatite in situ. Here, we studied and report the synthesis of nanohydroxyapatite and titanate nanotube (nHAp/TiNT) composites formulated at different concentrations (1, 2, 3, and 10 wt % TiNT) by means of a wet aqueous chemical reaction. The addition of TiNT affects the morphology of the nanocomposites, decreasing the average crystallite size from 54 nm (nHAp) to 34 nm (nHAp/TiNT10%), while confirming its interaction with the nanocomposite. The crystallinity index (CI) calculated by Raman spectroscopy and XRD showed that the values decreased according to the increase in TiNT concentration, which confirmed their addition to the structure of the nanocomposite. SEM images showed the presence of TiNTs in the nanocomposite. We further verified the potential cytotoxicity of murine fibroblast cell line L929, revealing that there was no remarkable cell death at any of the concentrations tested. In vivo regenerative activity was performed using oophorectomized animal (rat) models organized into seven groups containing five animals each over two experimental periods (15 and 30 days), with bone regeneration occurring in all groups tested within 30 days; however, the nHAp/TiNT10% group showed statistically greater tissue repair, compared to the untreated control group. Thus, the results of this study demonstrate that the presently formulated nHAp/TiNT nanocomposites are promising for numerous improved bone tissue regeneration applications.

Unraveling water monitoring association towards weather attributes for response proportions data: A unit-Lindley learning.

Learning techniques involve unraveling regression structures, which aim to analyze in a probabilistic frame the associations across variables of interest. Thus, analyzing fraction and/or proportion data may not be adequate with standard regression procedures, since the linear regression models generally assume that the dependent (outcome) variable is normally distributed. In this manner, we propose a statistical model called unit-Lindley regression model, for the purpose of Statistical Process Control (SPC). As a result, a new control chart tool was proposed, which targets the water monitoring dynamic, as well as the monitoring of relative humidity, per minute, of Copiapó city, located in Atacama Desert (one of the driest non-polar places on Earth), north of Chile. Our results show that variables such as wind speed, 24-hour temperature variation, and solar radiation are useful to describe the amount of relative humidity in the air. Additionally, Information Visualization (InfoVis) tools help to understand the time seasonality of the water particle phenomenon of the region in near real-time analysis. The developed methodology also helps to label unusual events, such as Camanchaca, and other water monitoring-related events.

An FDA-Approved Antifungal, Ketoconazole, and Its Novel Derivative Suppress tGLI1-Mediated Breast Cancer Brain Metastasis by Inhibiting the DNA-Binding Activity of Brain Metastasis-Promoting Transcription Factor tGLI1.

The goal of this study is to identify pharmacological inhibitors that target a recently identified novel mediator of breast cancer brain metastasis (BCBM), truncated glioma-associated oncogene homolog 1 (tGLI1). Inhibitors of tGLI1 are not yet available. To identify compounds that selectively kill tGLI1-expressing breast cancer, we screened 1527 compounds using two sets of isogenic breast cancer and brain-tropic breast cancer cell lines engineered to stably express the control, GLI1, or tGLI1 vector, and identified the FDA-approved antifungal ketoconazole (KCZ) to selectively target tGLI1-positive breast cancer cells and breast cancer stem cells, but not tGLI1-negative breast cancer and normal cells. KCZ's effects are dependent on tGLI1. Two experimental mouse metastasis studies have demonstrated that systemic KCZ administration prevented the preferential brain metastasis of tGLI1-positive breast cancer and suppressed the progression of established tGLI1-positive BCBM without liver toxicities. We further developed six KCZ derivatives, two of which (KCZ-5 and KCZ-7) retained tGLI1-selectivity in vitro. KCZ-7 exhibited higher blood-brain barrier penetration than KCZ/KCZ-5 and more effectively reduced the BCBM frequency. In contrast, itraconazole, another FDA-approved antifungal, failed to suppress BCBM. The mechanistic studies suggest that KCZ and KCZ-7 inhibit tGLI1's ability to bind to DNA, activate its target stemness genes Nanog and OCT4, and promote tumor proliferation and angiogenesis. Our study establishes the rationale for using KCZ and KCZ-7 for treating and preventing BCBM and identifies their mechanism of action.

Establishing a national reference laboratory for antimicrobial resistance using a whole-genome sequencing framework: Nigeria's experience.

Whole-genome sequencing (WGS) is finding important applications in the surveillance of antimicrobial resistance (AMR), providing the most granular data and broadening the scope of niches and locations that can be surveilled. A common but often overlooked application of WGS is to replace or augment reference laboratory services for AMR surveillance. WGS has supplanted traditional strain subtyping in many comprehensive reference laboratories and is now the gold standard for rapidly ruling isolates into or out of suspected outbreak clusters. These and other properties give WGS the potential to serve in AMR reference functioning where a reference laboratory did not hitherto exist. In this perspective, we describe how we have employed a WGS approach, and an academic-public health system collaboration, to provide AMR reference laboratory services in Nigeria, as a model for leapfrogging to national AMR surveillance.

Genomic characterization of invasive typhoidal and non-typhoidal Salmonella in southwestern Nigeria.

BACKGROUND: Salmonellosis causes significant morbidity and mortality in Africa. Information on lineages of invasive Salmonella circulating in Nigeria is sparse. METHODS: Salmonella enterica isolated from blood (n = 60) and cerebrospinal fluid (CSF, n = 3) between 2016 and 2020 from five tertiary hospitals in southwest Nigeria were antimicrobial susceptibility-tested and Illumina-sequenced. Genomes were analysed using publicly-available bioinformatic tools. RESULTS: Isolates and sequence types (STs) from blood were S. Typhi [ST1, n = 1 and ST2, n = 43] and invasive non-typhoidal Salmonella (iNTS) (S. Enteritidis [ST11, n = 7], S. Durham [ST10, n = 2], S. Rissen [ST8756, n = 2], S. Chester [ST2063, n = 1], S. Dublin [ST10, n = 1], S. Infantis [ST603, n = 1], S. Telelkebir [ST8757, n = 1] and S. Typhimurium [ST313, n = 1]). S. Typhi ST2 (n = 2) and S. Adabraka ST8757 (n = 1) were recovered from CSF. Most S. Typhi belonged to genotype 3.1.1 (n = 44), carried an IncY plasmid, had several antibiotic resistance genes (ARGs) including blaTEM-1 (n = 38), aph(6)-Id (n = 32), tet(A) (n = 33), sul2 (n = 32), dfrA14 (n = 30) as well as quinolone resistance-conferring gyrA_S83Y single-nucleotide polymorphisms (n = 37). All S. Enteritidis harboured aph(3")-Ib, blaTEM-1, catA1, dfrA7, sul1, sul2, tet(B) genes, and a single ARG, qnrB19, was detected in S. Telelkebir. Typhoidal toxins cdtB, pltA and pltB were detected in S. Typhi, Rissen, Chester, and Telelkebir. CONCLUSION: Most invasive salmonelloses in southwest Nigeria are vaccine-preventable infections due to multidrug-resistant, West African dominant S. Typhi lineage 3.1.1. Invasive NTS serovars, including some harbouring typhoidal toxin or resistance genes, represented a third of the isolates emphasizing the need for better diagnosis and surveillance.

Ionizable Lipid Nanoparticle-Mediated Delivery of Plasmid DNA in Cardiomyocytes.

Introduction: Gene therapy is a promising approach to be applied in cardiac regeneration after myocardial infarction and gene correction for inherited cardiomyopathies. However, cardiomyocytes are crucial cell types that are considered hard-to-transfect. The entrapment of nucleic acids in non-viral vectors, such as lipid nanoparticles (LNPs), is an attractive approach for safe and effective delivery. Methods: Here, a mini-library of engineered LNPs was developed for pDNA delivery in cardiomyocytes. LNPs were characterized and screened for pDNA delivery in cardiomyocytes and identified a lead LNP formulation with enhanced transfection efficiency. Results: By varying lipid molar ratios, the LNP formulation was optimized to deliver pDNA in cardiomyocytes with enhanced gene expression in vitro and in vivo, with negligible toxicity. In vitro, our lead LNP was able to reach a gene expression greater than 80%. The in vivo treatment with lead LNPs induced a twofold increase in GFP expression in heart tissue compared to control. In addition, levels of circulating myeloid cells and inflammatory cytokines remained without significant changes in the heart after LNP treatment. It was also demonstrated that cardiac cell function was not affected after LNP treatment. Conclusion: Collectively, our results highlight the potential of LNPs as an efficient delivery vector for pDNA to cardiomyocytes. This study suggests that LNPs hold promise to improve gene therapy for treatment of cardiovascular disease.