Transporter Gene Research Articles

Use of transcriptomics and genomics to assess the effect of disinfectant exposure on the survival and resistance of Escherichia coli O157:H7, a human pathogen.

Disinfectants are essential for biosecurity, preventing the persistence and spread of zoonotic pathogens on farms and subsequent human infections. In this study, transcriptomics and genomics were utilised to assess the effect of disinfectant exposure on pathogenic Escherichia coli. The exposure of E. coli O157:H7 to sub-optimal concentrations of commonly used farm disinfectants elicited changes in both the transcriptome and genome. The transcriptomics identified upregulation of >300 genes and downregulation of >100 genes with functions, which included stress response, metabolism, transcription, transportation, membrane-associated and virulence genes. The phage shock protein (psp) operon was highly upregulated in response to a quaternary ammonium compound (QAC)-containing disinfectant, which has not previously been associated with a response to chemical stress. Disinfectant-adapted isolates generated by exposure to sub-lethal disinfectants levels demonstrated resistance to several common antibiotics and decreased sensitivity to biocides. Whole genome sequencing of the mutant strains indicated that they had acquired mutations in the genes associated with the upregulation of the multiple antibiotic resistance (MAR) efflux system (lon protease and marR) and topoisomerase genes (gyrA and gyrB). The disinfectant-adapted isolates also exhibited increased expression of transcription, respiration and several pH stress response genes localised in the "acid fitness island." This study demonstrated that sub-optimal disinfectant concentrations allow E. coli O157:H7 to adapt and survive disinfection and develop antibiotic resistance. These changes could have implications for disease treatment and elimination on farms. Although E. coli O157:H7 and farm disinfectants were the focus of this study, we believe these findings are also applicable to other settings, including hospitals.

Engineering Yarrowia lipolytica to Produce l-Malic Acid from Glycerol.

The declining availability of cheap fossil-based resources has sparked growing interest in the sustainable biosynthesis of organic acids. l-Malic acid, a crucial four-carbon dicarboxylic acid, finds extensive applications in the food, chemical, and pharmaceutical industries. Synthetic biology and metabolic engineering have enabled the efficient microbial production of l-malic acid, albeit not in Yarrowia lipolytica, an important industrial microorganism. The present study aimed to explore the potential of this fungal species for the production of l-malic acid. First, endogenous biosynthetic genes and heterologous transporter genes were overexpressed in Y. lipolytica to identify bottlenecks in the l-malic acid biosynthesis pathway grown on glycerol. Second, overexpression of isocitrate lyase, malate synthase, and malate dehydrogenase in the glyoxylate cycle pathway and introduction of a malate transporter from Schizosaccharomyces pombe significantly boosted l-malic acid production, which reached 27.0 g/L. A subsequent increase to 37.0 g/L was attained through shake flask medium optimization. Third, adaptive laboratory evolution allowed the engineered strain Po1g-CEE2+Sp to tolerate a lower pH and to accumulate a higher amount of l-malic acid (56.0 g/L). Finally, when scaling up to a 5 L bioreactor, a titer of 112.5 g/L was attained. In conclusion, this study demonstrates for the first time the successful production of l-malic acid in Y. lipolytica by combining metabolic engineering and laboratory evolution, paving the way for large-scale sustainable biosynthesis of this and other organic acids.

Trehalose metabolism mediates trade-offs between reproduction and survival in beet webworm, Loxostege sticticalis, under heat stress.

Temperature is an important determinant of developmental and reproductive rates in insects. Here, we investigated the physiological responses of adult beet webworm, Loxostege sticticalis L. (Lepidoptera: Crambidae), to three temperatures (16, 23 and 30 °C) focusing on trehalose metabolism. Exposure of moths to 30 °C accelerated eclosion and ovarian development, but shortened the oviposition period and adult longevity, whereas exposure to 16 °C had opposite effects. Transcriptome analysis revealed that vitellogenin (VG) and vitellogenin receptor (VR) genes were up-regulated at 30 °C, as were numerous genes related to energy metabolism, including those involved in the insulin signaling pathway, the tricarboxylic acid (TCA) cycle, and glycolysis. Expression of the trehalose transporter gene TRET1 was also induced at high temperature, primarily in the ovaries, where trehalose content increased, accompanied by lipid degradation in the fat body. Treatment with the trehalase inhibitor validamycin A reduced female fecundity and longevity at 23 °C, but enhanced the expression of genes related to stress resistance and reproduction, mimicking the effect of high temperature. Besides their practical utility for predicting the oviposition behavior and geographic distribution of L. sticticalis in the field, these results elucidate the various physiological roles of trehalose in L. sticticalis during exposure of moths to high temperature and may provide insights into the relationship between stress resistance and reproduction in insects more generally. © 2024 Society of Chemical Industry.

Interaction of Val66Met Brain-Derived Neurotrophic Factor and 5-HTTLPR Serotonin Transporter Gene Polymorphisms with Lifetime Prevalence of Post-Traumatic Stress Disorder in Primary Care Patients

Background/Objectives: Post-traumatic stress disorder (PTSD) is a complex condition influenced by both genetic and environmental factors. This longitudinal study aimed to explore the connection between two specific genetic polymorphisms, Val66Met and 5-HTTLPR, and the lifetime prevalence of PTSD in patients from primary care settings. We also examined the role of sociodemographic and psychosocial factors to provide a more comprehensive view of PTSD risk. Methods: We recruited a cohort of primary care patients and diagnosed PTSD using a standardized diagnostic interview. Genetic analyses focused on Val66Met and 5-HTTLPR polymorphisms. We applied logistic regression to assess the association between these genetic markers and PTSD, considering factors such as gender, family history of depression, and experiences of childhood maltreatment. Results: Our findings show that women, individuals with a family history of depression, and those exposed to childhood maltreatment have a higher risk of developing PTSD. While the Val66Met polymorphism was not significantly associated with PTSD, the 5-HTTLPR polymorphism showed a marginal relationship. No significant interaction was found between the two polymorphisms in relation to PTSD. Conclusions: This study underscores the multifactorial nature of PTSD, influenced by both genetic and environmental factors. The findings point to the importance of further research on genetic predispositions and highlight the value of early interventions for high-risk populations in primary care settings.

Structural Derivatives of β-Asarone from Acorus calamus Linn. as Insecticide Candidates and the Insecticidal Mechanism Against Small Brown Planthopper

The small brown planthopper (SBPH), Laodelphax striatellus (Fallén) (Hemiptera: Delphacidae), is an increasing threat to Gramineae crops, posing significant risks to both the environment and food safety. β-asarone, as a promising green alternative to chemical insecticides, possesses wide application prospects in the crop protection field. To enhance the insecticidal activity of β-asarone, a series of derivatives were prepared through an active substructure splicing strategy, and their insecticidal activities against SBPH were evaluated. Among the 7 commercial compounds with chemical structures similar to β-asarone and 12 structural derivatives of β-asarone, compound 10, which incorporates the 2-chloropyridine functional group from flupyrimin, exhibited the most potent insecticidal activity against SBPH, with an 8.31-fold increase in insecticidal activity compared to β-asarone. Furthermore, transcriptome analysis showed that among the selected genes that may play important roles in insecticidal activity, an ABC transporter gene, MDR49, was most significantly down-regulated. MDR49 was highly expressed in the 4th-instar nymphs, with the highest expression level in the fat body, midgut, and abdomen. RNA interference (RNAi) against MDR49 significantly reduced susceptibility to compound 10 in SBPH, which revealed that MDR49 may be the candidate insecticidal target of compound 10. Additionally, the insecticidal spectrum revealed that compound 10 showed excellent efficacy against Bemisia tabaci (Gennadius) (Hemiptera: Aleyrodidae) and Tetranychus cinnabarinus (Boisduval) (Acarina: Tetranychidae). This study indicates that compound 10 could be further developed as a novel eco-friendly pesticide.

The mayfly Neocloeon triangulifer senses decreasing oxygen availability (PO2) and responds by reducing ion uptake and altering gene expression.

Oxygen availability is central to the energetic budget of aquatic animals and may vary naturally and/or in response to anthropogenic activities. Yet, we know little about how oxygen availability is linked to fundamental processes such as ion transport in aquatic insects. We hypothesized and observed that ion (22Na and 35SO4) uptake would be significantly decreased at O2 partial pressures below the mean Pcrit (5.4 kPa) where metabolic rates (MO2) are compromised, and ATP production is limited. However, we were surprised to observe marked reductions in ion uptake at oxygen partial pressures well above the Pcrit, where MO2 was stable. For example, SO4 uptake decreased by 51% at 11.7kPa, and 82% at the Pcrit (5.4kPa) while Na uptake decreased by 19% at 11.7kPa, and 60% at the Pcrit. Nymphs held for longer time periods at reduced PO2 exhibited stronger reductions in ion uptake rates. Fluids from whole body homogenates exhibited a 29% decrease in osmolality in the most hypoxic condition. The differential expression of atypical guanylyl cyclase (gcy-88e) in response to changing PO2 conditions provides evidence for its potential role as an oxygen sensor. Several ion transport genes (e.g., chloride channel and sodium-potassium ATPase) and hypoxia-associated genes (e.g., ldh and egl-9) were also impacted by decreased oxygen availability. Together, our work suggests that N. triangulifer can sense decreased oxygen availability and perhaps conserves energy accordingly, even when MO2 is not impacted.

Hyperosmotic Stress Induces the Expression of Organic Osmolyte Transporters in Porcine Intestinal Cells and Betaine Exerts a Protective Effect on the Barrier Function.

Background/objectives: The porcine intestinal epithelium plays a fundamental role as a defence interface against pathogens. Its alteration can cause severe inflammatory conditions and diseases. Hyperosmotic stress under physiological conditions and upon pathogen challenge can cause malabsorption. Different cell types counteract the osmolarity increase by accumulating organic osmolytes such as betaine, taurine, and myo-inositol through specific transporters. Betaine is known for protecting cells from hyperosmotic stress and has positive effects when fed to pigs. The aim of this study is to demonstrate the modulation of osmolyte transporters gene expression in IPEC-J2 during osmolarity changes and assess the effects of betaine. Methods: IPEC-J2 were seeded in transwells, where differentiate as a polarized monolayer. Epithelial cell integrity (TEER), oxidative stress (NO) and gene expression of osmolyte transporters, tight junction proteins (TJp) and pro-inflammatory cytokines were evaluated. Results: Cells treated with NaCl hyperosmolar medium (500 mOsm/L) showed a TEER decrease at 3 h and detachment within 24 h, associated with an osmolyte transporters reduction. IPEC-J2 treated with mannitol hyperosmolar medium (500 mOsm/L) upregulated taurine (TauT), myo-inositol (SMIT) and betaine (BGT1) transporters expression. A decrease in TJp expression was associated with a TEER decrease and an increase in TNFα, IL6, and IL8. Betaine could attenuate the hyperosmolarity-induced reduction in TEER and TJp expression, the NO increase and cytokines upregulation. Conclusions: This study demonstrates the expression of osmolyte transporters in IPEC-J2, which was upregulated upon hyperosmotic treatment. Betaine counteracts changes in intracellular osmolarity by contributing to maintaining the epithelial barrier function and reducing the inflammatory condition. Compatible osmolytes may provide beneficial effects in therapies for diseases characterized by inflammation and TJp-related dysfunctions.

Diverse and specialized metabolic capabilities of microbes in oligotrophic built environments

BackgroundBuilt environments (BEs) are typically considered to be oligotrophic and harsh environments for microbial communities under normal, non-damp conditions. However, the metabolic functions of microbial inhabitants in BEs remain poorly understood. This study aimed to shed light on the functional capabilities of microbes in BEs by analyzing 860 representative metagenome-assembled genomes (rMAGs) reconstructed from 738 samples collected from BEs across the city of Hong Kong and from the skin surfaces of human occupants. The study specifically focused on the metabolic functions of rMAGs that are either phylogenetically novel or prevalent in BEs.ResultsThe diversity and composition of BE microbiomes were primarily shaped by the sample type, with Micrococcus luteus and Cutibacterium acnes being prevalent. The metabolic functions of rMAGs varied significantly based on taxonomy, even at the strain level. A novel strain affiliated with the Candidatus class Xenobia in the Candidatus phylum Eremiobacterota and two novel strains affiliated with the superphylum Patescibacteria exhibited unique functions compared with their close relatives, potentially aiding their survival in BEs and on human skins. The novel strains in the class Xenobia possessed genes for transporting nitrate and nitrite as nitrogen sources and nitrosative stress mitigation induced by nitric oxide during denitrification. The two novel Patescibacteria strains both possessed a broad array of genes for amino acid and trace element transport, while one of them carried genes for carotenoid and ubiquinone biosynthesis. The globally prevalent M. luteus in BEs displayed a large and open pangenome, with high infraspecific genomic diversity contributed by 11 conspecific strains recovered from BEs in a single geographic region. The versatile metabolic functions encoded in the large accessory genomes of M. luteus may contribute to its global ubiquity and specialization in BEs.ConclusionsThis study illustrates that the microbial inhabitants of BEs possess metabolic potentials that enable them to tolerate and counter different biotic and abiotic conditions. Additionally, these microbes can efficiently utilize various limited residual resources from occupant activities, potentially enhancing their survival and persistence within BEs. A better understanding of the metabolic functions of BE microbes will ultimately facilitate the development of strategies to create a healthy indoor microbiome.Bh9ATT9ssZEtNaJHv1wg7GVideo

Inorganic nitrogen inhibits symbiotic nitrogen fixation through blocking NRAMP2-mediated iron delivery in soybean nodules

Symbiotic nitrogen fixation (SNF) in legume-rhizobia serves as a sustainable source of nitrogen (N) in agriculture. However, the addition of inorganic N fertilizers significantly inhibits SNF, and the underlying mechanisms remain not-well understood. Here, we report that inorganic N disrupts iron (Fe) homeostasis in soybean nodules, leading to a decrease in SNF efficiency. This disruption is attributed to the inhibition of the Fe transporter genes Natural Resistance-Associated Macrophage Protein 2a and 2b (GmNRAMP2a&2b) by inorganic N. GmNRAMP2a&2b are predominantly localized at the tonoplast of uninfected nodule tissues, affecting Fe transfer to infected cells and consequently, modulating SNF efficiency. In addition, we identified a pair of N-signal regulators, nitrogen-regulated GARP-type transcription factors 1a and 1b (GmNIGT1a&1b), that negatively regulate the expression of GmNRAMP2a&2b, which establishes a link between N signaling and Fe homeostasis in nodules. Our findings reveal a plausible mechanism by which soybean adjusts SNF efficiency through Fe allocation in response to fluctuating inorganic N conditions, offering valuable insights for optimizing N and Fe management in legume-based agricultural systems.

Exome Sequence Data of Eight SLC Transporters Reveal That SLC22A1 and SLC22A3 Variants Alter Metformin Pharmacokinetics and Glycemic Control.

Background: Type 2 diabetes (T2D) is one of the leading causes of mortality and is a public health challenge worldwide. Metformin is the first-choice treatment for T2D; its pharmacokinetics (PK) is facilitated by members of the solute carrier (SLC) superfamily of transporters, it is not metabolized, and it is excreted by the kidney. Although interindividual variability in metformin pharmacokinetics is documented in the Mexican population, its pharmacogenomics is still underexplored. We aimed to identify variants in metformin SLC transporter genes associated with metformin PK and response in Mexican patients. Methods: Using exome data from 2217 Mexican adults, we identified 86 biallelic SNVs in the eight known genes encoding SLC transporters, with a minor allele frequency ≥ 1%, which were analyzed in an inadequate glycemic control (IGC) association study in T2D metformin treated patients. Metformin PK was evaluated in a pediatric cohort and the effect of associated SNVs was correlated. Results: Functional annotation classified two SNVs as pathogenic. The association study revealed two blocks associated with IGC. These haplotypes comprise rs622591, rs4646272, rs4646273, and rs4646276 in SLC22A1; and rs1810126 and rs668871 in SLC22A3. PK profiles revealed that homozygotes of the SLC22A1 haplotype reached lower plasma metformin concentrations 2 h post administration than the other groups. Conclusions: Our findings highlight the potential of pharmacogenomics studies to enhance precision medicine, which may involve dosage adjustments or the exploration of alternative therapeutic options. These hold significant implications for public health, particularly in populations with a high susceptibility to develop metabolic diseases, such as Latin Americans.

Lactobacillus gasseri BNR17 and Limosilactobacillus fermentum ABF21069 Ameliorate High Sucrose-Induced Obesity and Fatty Liver via Exopolysaccharide Production and β-oxidation.

Obesity and metabolic dysfunction-associated fatty liver disease (MAFLD) are prevalent metabolic disorders with substantial global health implications that are often inadequately addressed by current treatments and may have side effects. Probiotics have emerged as promising therapeutic agents owing to their beneficial effects on gut health and metabolism. This study investigated the synergistic effects of a probiotic combination of BNR17 and ABF21069 on obesity and MAFLD in C57BL/6 mice fed a high-sucrose diet. The probiotic combination significantly reduced body weight and fat accumulation compared with the high-sucrose diet. It also alleviated elevated serum leptin levels induced by a high-sucrose diet. Histological analysis revealed a significant reduction in white adipose tissue and fatty liver in the mice treated with the probiotic combination. Furthermore, increased expression of genes related to β-oxidation, thermogenesis, and lipolysis suggested enhanced metabolic activity. The probiotic groups, particularly the BNR17 group, showed an increase in fecal exopolysaccharides, along with a tendency toward a lower expression of intestinal sugar transport genes, indicating reduced sugar absorption. Additionally, inflammatory markers in the liver tissue exhibited lower expression in the ABF21069 group than in the HSD group. Despite each strain in the combination group having distinct characteristics and functions, their combined effect demonstrated synergy in mitigating obesity and MAFLD, likely through the modulation of fecal exopolysaccharides content and improvement in lipid metabolism. These findings underscore the potential of probiotic supplementation as a promising assistant therapy for managing obesity and MAFLD and provide valuable insights into its therapeutic mechanisms in metabolic disorders.

Chitosan nanoparticles (CSNPs) conferred salinity tolerance in maize by upregulating E3 ubiquitin-protein ligase, P5CS1, HKT1, NHX1, and PMP3 genes.

This study explored the transcriptional behaviors of several candidate genes in response to the application of CSNPs (50 and 100 mgl-1) in maize seedlings grown under two salinity levels (NaCl of 0.07 and 0.14 gkg-1soil). Employing CSNPs at both concentrations mitigated the inhibitory role of salinity on the leaf and root fresh weights. The application of CSNPs enhanced the transcription of the E3 ubiquitin-protein ligase gene by an average of threefold, contrasted with the salinity controls. The Δ1-pyrroline-5-carboxylate synthetase (P5CS1) gene was upregulated in response to both individual and mixed treatments of CSNPs and salinity. The transcription of the high-affinity K+ transporter (HKT1) gene displayed an upward trend in response to the CSNPs and salinity treatments. The Na+/H+ exchangers (NHX1) gene exhibited a similar trend to that of the HKT1 gene. The utilization of CSNPs was accompanied by an upregulation in the plasma membrane proteolipid 3 (PMP3) gene, contrasted with the salinity controls. The phenylalanine ammonia-lyase (PAL) activity displayed an upward trend in response to the foliar application of CSNPs. The CSNPs at the 100 mgl-1 concentration were more capable of inducing the ascorbate peroxidase enzyme under both salinity conditions than the 50 mgl-1 dose. The simultaneous exposure of maize seedlings to CSNPs and salinity resulted in the drastic upregulation of the catalase activities. This study provides novel insights into the major mechanisms underlying the stress-mitigating effects of CSNPs, thereby providing a suitable platform for their application in sustainable agricultural practices.

Growth stage-related capsular polysaccharide translocon Wza in Vibrio splendidus modifies phage vB_VspM_VS2 susceptibility

Bacteria at different growth stages usually coordinate capsular polysaccharide (CPS) formation and may affect their susceptibility to phage. In this study, we evaluated the infection efficacy of phage vB_VspM_VS2 in V. splendidus AJ01 at different growth stages and explored the role of growth stage-related CPS translocon Wza in the susceptibility of V. splendidus to phage vB_VspM_VS2. The results showed that V. splendidus locked in the stationary growth stage (SGS) or early exponential stage (EES) infected with phage (EES-P) has a low susceptibility to phage vB_VspM_VS and exhibit a pronounced reduction in phage adsorption rate as compared to the EES bacteria. The expression of wza of CPS transport gene was significantly increased in EES bacteria compared to that bacteria locked in the SGS or EES-P. Bacteria with deleted wza (Δwza mutant) escaped phage adsorption due to absence of Wza mediated down-regulation of CPS expression, otherwise. Our results reveal that the Wza of V. splendidus can promotes phage to infect these bacteria via increasing the phage absorption, which provides important implications for using phages therapeutically target pathogenic bacteria in dynamics communities.

Novel NhaC Na+/H+ antiporter in cyanobacteria contributes to key molecular processes for salt tolerance.

Genome mining has revealed the halotolerant cyanobacterium Halothece sp. PCC7418 harbors considerable enrichment in the ion transport gene family for putative Na+/H+ antiporters. Here, we compared transcriptomic profiles of these encoding genes under various abiotic stresses and discovered that Halothece NhaC (hnhaC) was one of 24 genes drastically upregulated under salt stress. Critical roles of HnhaC in salt-stress protection and response were identified by a complementation assay using the salt-sensitive mutant Escherichia coli strain TO114. Expression of HnhaC rendered this mutant more tolerant to high concentrations of NaCl and LiCl. Antiporter activity assays showed that HnhaC protein predominantly exhibited Na+/H+ and Li+/H+ antiporter activities under neutral or alkaline pH conditions. Furthermore, expression of HnhaC conferred adaptive benefits onto E. coli by enabling a conditional filamentation phenotype. Dissecting the molecular mechanism of this phenotype revealed that differentially expressed genes were associated with clusters of SOS-cell division inhibitor, SOS response repair, and Z-associated proteins. Together, these results strongly indicate that HnhaC is an Na+/H+ antiporter that contributes to salt tolerance. The ubiquitous existence of several Na+/H+ antiporters represents a complex molecular system in halotolerant cyanobacteria, which can be deployed differently in response to growth and to environmental stresses.

Expression of the grapevine anion transporter ALMT2 in Arabidopsis root decreases shoot Cl-/NO3- ratio under salt stress.

Grapevines (Vitis vinifera, Vvi) are economically important crop plants which, when challenged with salt (NaCl) in soil and/or irrigation water, tend to accumulate Na+ and Cl- in aerial tissues impacting yield, and berry acceptability for winemaking. Grapevine (Vitis spp.) rootstocks vary in their capacity for shoot Cl- exclusion. Here, we characterise two putative anion transporter genes - Aluminium-activated Malate Transporter VviALMT2 and VviALMT8 - that were differentially expressed in the roots of efficient (140 Ruggeri) and inefficient (K51-40) Cl- excluding rootstocks, to explore their potential for impacting shoot Cl- exclusion. Using the Xenopus laevis oocyte expression system, VviALMT2 and VviALMT8 formed conductive channels that were highly permeable to NO3-, slightly-to-moderately permeable to other substrates including Cl- and malate, but impermeable to SO42-. RT-qPCR analyses revealed that VviALMT2 was more highly expressed in the root vasculature and up-regulated by high [NO3-] re-supply post starvation, while fluorescently tagged translational fusion VviALMT2 localised to the plasma membrane. As VviALMT8 showed no such features, we selected VviALMT2 as our salt exclusion candidate and assessed its function in planta. Expression of VviALMT2 in Arabidopsis thaliana root vasculature reduced shoot [Cl-]/[NO3-] after NaCl treatment, which suggests that VviALMT2 can be beneficial to plants under salt stress.

Impact of early postbiotic supplementation on broilers’ responses to subclinical necrotic enteritis

Necrotic enteritis (NE), an enteric disease caused by Clostridium perfringens, results in damage to the intestinal epithelial lining disrupting its function, nutrient absorption, and utilization. This study evaluated the effects of in ovo and post-hatch applications of a Saccharomyces cerevisiae-based postbiotic on performance and nutrient transporter genes of broilers during a NE challenge. At embryonic d 18, Ross 708 fertile eggs were injected with 0.2 mL of either water or postbiotic. A total of 288 male hatchlings were assigned to one of the following treatment groups: 1) NC (in ovo water injection, no challenge); 2) PIW (postbiotic in ovo and in drinking water, no challenge); 3) NC+ (NC with NE challenge); and 4) PIW+ (PIW with NE challenge). On d 14, all birds in the NE-challenged groups were orally gavaged with 3,000 Eimeria maxima sporulated oocysts followed by two doses of ∼1×108 CFU/mL/bird of C. perfringens on d 19 and d 20. Hatchability, weekly performance, intestinal lesion scores, and mRNA abundance of nutrient transporters in the jejunum and ileum were assessed. Data were analyzed by 2-way ANOVA in JMP and significance between treatments identified by LSD test (P ≤ 0.05). A significant postbiotic treatment and NE challenge interaction was observed in performance during d 21-28 with a greater ADG in PIW compared to NC and PIW+. Lesion scores in the jejunum and ileum were significantly reduced in PIW+ compared to NC+. On d 7, mRNA abundance of SGLT1 was significantly greater in PIW compared to the NC group. On d 14, birds in PIW had greater levels of GLUT2 and EAAT3 than NC group. No significant interaction effects were observed on d 21. PIW+ had significantly greater EAAT3 mRNA levels compared to PIW in jejunum and PIW and NC+ in ileum on d 28. In conclusion, in ovo and water supplementation of this postbiotic presents a potential to improve the performance, ameliorate pathology detriments associated with NE, and positively regulate the mRNA levels of key nutrient transporters during NE challenge.

Response to therapy of creatine transporter deficiency caused by a hypomorphic variant in SLC6A8

Cerebral creatine deficiency syndromes (CCDS) are rare inherited metabolic disorders caused by defective biosynthesis or transport of creatine. These conditions are characterized by reduced accumulation of creatine in the brain, mild to severe intellectual disability, global developmental delay, and speech-language disorders. The amount of brain creatine reduction needed to cause symptoms is not known. Here we report a new patient with creatine transporter deficiency (CTD) who presented at 15 months of age with seizures and global delays with no speech at 3 years of age. Brain MRI was normal, but brain MRS indicated creatine levels reduced to about 20 % of normal. He had normal levels of creatine and guanidinoacetate in plasma, but increased creatine/creatinine ratio in urine. DNA sequencing identified a hemizygous c.832C > T (p.Arg278Cys) variant in the creatine transporter gene SLC6A8. Fibroblasts from this patient had about 25 % of normal creatine transport activity, a value much higher than that measured in patients whose variants introduced premature stop codons in SLC6A8. The child was started on supplements of creatine, glycine, and arginine. His speech improved dramatically, and he had no more seizures, even during episodes of fever. Despite the clinical improvement, a repeat MRS demonstrated similar levels of brain creatine. This study suggests that a reduction in creatine transporter activity to 25 % or less is sufficient to cause symptoms of brain creatine deficiency and that functionally milder forms of CTD might respond to supplements aimed at replenishing brain creatine.

IPSC-derived blood-brain barrier modeling reveals APOE isoform-dependent interactions with amyloid beta

BackgroundThree common isoforms of the apolipoprotein E (APOE) gene - APOE2, APOE3, and APOE4 - hold varying significance in Alzheimer’s Disease (AD) risk. The APOE4 allele is the strongest known genetic risk factor for late-onset Alzheimer’s Disease (AD), and its expression has been shown to correlate with increased central nervous system (CNS) amyloid deposition and accelerated neurodegeneration. Conversely, APOE2 is associated with reduced AD risk and lower CNS amyloid burden. Recent clinical data have suggested that increased blood-brain barrier (BBB) leakage is commonly observed among AD patients and APOE4 carriers. However, it remains unclear how different APOE isoforms may impact AD-related pathologies at the BBB.MethodsTo explore potential impacts of APOE genotypes on BBB properties and BBB interactions with amyloid beta, we differentiated isogenic human induced pluripotent stem cell (iPSC) lines with different APOE genotypes into both brain microvascular endothelial cell-like cells (BMEC-like cells) and brain pericyte-like cells. We then compared the effect of different APOE isoforms on BBB-related and AD-related phenotypes. Statistical significance was determined via ANOVA with Tukey’s post hoc testing as appropriate.ResultsIsogenic BMEC-like cells with different APOE genotypes had similar trans-endothelial electrical resistance, tight junction integrity and efflux transporter gene expression. However, recombinant APOE4 protein significantly impeded the “brain-to-blood” amyloid beta 1–40 (Aβ40) transport capabilities of BMEC-like cells, suggesting a role in diminished amyloid clearance. Conversely, APOE2 increased amyloid beta 1–42 (Aβ42) transport in the model. Furthermore, we demonstrated that APOE-mediated amyloid transport by BMEC-like cells is dependent on LRP1 and p-glycoprotein pathways, mirroring in vivo findings. Pericyte-like cells exhibited similar APOE secretion levels across genotypes, yet APOE4 pericyte-like cells showed heightened extracellular amyloid deposition, while APOE2 pericyte-like cells displayed the least amyloid deposition, an observation in line with vascular pathologies in AD patients.ConclusionsWhile APOE genotype did not directly impact general BMEC or pericyte properties, APOE4 exacerbated amyloid clearance and deposition at the model BBB. Conversely, APOE2 demonstrated a potentially protective role by increasing amyloid transport and decreasing deposition. Our findings highlight that iPSC-derived BBB models can potentially capture amyloid pathologies at the BBB, motivating further development of such in vitro models in AD modeling and drug development.

Dopamine Transporter and CYP2D6 Gene Relationships with Attention-Deficit/Hyperactivity Disorder Treatment Response in the Methylphenidate and Atomoxetine Crossover Study.

Background: Few biological or clinical predictors guide medication selection and/or dosing for attention-deficit/hyperactivity disorder (ADHD). Accumulating data suggest that genetic factors may contribute to clinically relevant pharmacodynamic (e.g., dopamine transporter-SLC6A3 also commonly known as DAT1) or pharmacokinetic (e.g., the drug metabolizing enzyme Cytochrome P450 2D6 CYP2D6) effects of methylphenidate (stimulant) and atomoxetine (non-stimulant), which are commonly prescribed medications. This is the first study of youth with ADHD exposed to both medications examining the clinical relevance of genetic variation on treatment response. Methods: Genetic variations in DAT1 and CYP2D6 were examined to determine how they modified time relationships with changes in ADHD symptoms over a 4-week period in 199 youth participating in a double-blind crossover study following a stepped titration dose optimization protocol. Results: Our results identified trends in the modification effect from CYP2D6 phenotype and the time-response relationship between ADHD total symptoms for both medications (atomoxetine [ATX]: p = 0.058, Methylphenidate [MPH]: p = 0.044). There was also a trend for the DAT1 3' untranslated region (UTR) variable number of tandem repeat (VNTR) genotype to modify dose relationships with ADHD-RS total scores for atomoxetine (p = 0.029). Participants with DAT1 9/10 repeat genotypes had a more rapid dose-response to ATX compared to 10/10, while those with 9/9 genotypes did not respond as doses were increased. Regardless of genotype, ADHD symptoms and doses were similar across CYP2D6 metabolizer groups after 4 weeks of treatment. Conclusions: Most children with ADHD who were CYP2D6 normal metabolizers or had DAT1 10/10 or 9/10 genotypes responded well to both medications. While we observed some statistically significant effects of CYP2D6 and DAT1 with treatment response over time, our data indicate that genotyping for clinical purposes may have limited utility to guide treatment decisions for ATX or MPH because both medications were generally effective in the studied cohort after 3 weeks of titration to higher doses. The potential DAT1 association with ATX treatment is a novel finding, consistent with prior reports suggesting an association of the DAT1 in 9/9 genotypes with lower responsive rates to treatment at low and moderate doses.

Unique rumen micromorphology and microbiota-metabolite interactions: features and strategies for Tibetan sheep adaptation to the plateau.

The rumen microbiota-a symbiont to its host and consists of critical functional substances-plays a vital role in the animal body and represents a new perspective in the study of adaptive evolution in animals. This study used Slide Viewer slicing analysis system, gas chromatography, RT-qPCR and other technologies, as well as 16S and metabolomics determination methods, to measure and analyze the microstructure of rumen epithelium, rumen fermentation parameters, rumen transport genes, rumen microbiota and metabolites in Tibetan sheep and Hu sheep. The results indicate that the rumen nipple height and cuticle thickness of Tibetan sheep are significantly greater than those of Hu sheep (p < 0.01) and that the digestion and absorption of forage are greater. The levels of carbohydrate metabolism, lipid metabolism, and protein turnover were increased in Tibetan sheep, which enabled them to ferment efficiently, utilize forage, and absorb metabolic volatile fatty acids (VFAs). Tibetan sheep rumen metabolites are related to immune function and energy metabolism, which regulate rumen growth and development and gastrointestinal homeostasis. Thus, compared with Hu sheep, Tibetan sheep have more rumen papilla and cuticle corneum, and the synergistic effect of the microbiota and its metabolites is a characteristic and strategy for adapting to high-altitude environments.