Altered Cell Permeability Research Articles

Single Nuclei Transcriptomics in Diabetic Mice Reveals Altered Brain Hippocampal Endothelial Cell Function, Permeability, and Behavior

Single Nuclei Transcriptomics in Diabetic Mice Reveals Altered Brain Hippocampal Endothelial Cell Function, Permeability, and Behavior

Single nuclei transcriptomics in diabetic mice reveals altered brain hippocampal endothelial cell function, permeability, and behavior

Type 2 diabetes mellitus (T2DM) is a metabolic disorder with cerebrovascular and cardiovascular sequelae. Yet, a clear pattern of gene dysregulation by T2DM in dementia has yet to be defined. We used single nuclei RNA sequencing technology to profile the transcriptome of endothelial cells (EC) from anatomically defined hippocampus of db/db mice to identify differentially expressed (DE) genes, gene pathways and networks, predicted regulating transcription factors, and targets of DE long noncoding RNAs. We also applied gadolinium (Gd) enhanced magnetic resonance imaging (MRI) to assess blood brain barrier (BBB) permeability, and functionally assessed cognitive behavior. The murine gene expression profiles were then integrated with those of persons with Alzheimer's disease (AD) and vascular dementia (VaD).We reveal that the transcriptome of the diabetic hippocampal murine brain endothelium differs substantially from control wild types with molecular changes characterized by differential RNA coding and noncoding pathways enriched for EC signaling and for endothelial functions for neuroinflammation, endothelial barrier disruption, and neurodegeneration. Gd enhanced structural brain MRI linked endothelial molecular alterations to BBB dysfunction by neuroimaging. Integrated multiomics of hippocampal endothelial gene dysregulation associated with impairments in cognitive adaptive capacity. In addition, the diabetic transcriptome significantly and positively correlated with that of persons with AD and VaD. Taken together, our results from comprehensive, multilevel, integrated, single nuclei transcriptomics support the hypothesis of T2DM-mediated neuroinflammation and endothelial cell and barrier disruption as key mechanisms in cognitive decline in T2DM, thereby suggesting potential endothelial-specific molecular therapeutic targets.

Alteration of Azole-Resistant Candida albicans Cell Permeability by a Flavonoid from Dalea elegans

Background: The flavonoid 2′,4′-dihydroxy-5′-(1′′′,1′′′-dimethylalil)-8-prenylpinocembrin (8PP), obtained from Dalea elegans roots has demonstrated antifungal activities. It also overcomes the resistance to azoles in azole-resistant Candida albicans (RCa). It collaborates with fluconazole to decrease cell growth and viability and inhibits ABC cdr transporters at the carrying site and associated ATPase. In addition, 8PP reduces cell viability dependent on oxidoreductase activity in planktonic cultures. It shows a dual oxidant-antioxidant activity with accumulating endogenous reactive oxygen species (ROS) in RCa biofilms. Objectives: This work evaluated the effects of 8PP, fluconazole, and their combination on RCa viability, which is dependent on cell permeability. Methods: RCa 12 - 99 strain of oral origin was used. Its viability was assessed through propidium iodide (PI) cell uptake and observing yeast morphology by fluorescence microscopy. Results: RCa 12 - 99 strain cells, collected in the mid-exponential phase, incorporated PI and completely stained red after incubation with 8PP or fluconazole at concentrations that inhibit cell growth and viability. When combined at 125 µM of each, 8PP and fluconazole appeared more effective than separately. The results indicated that the compounds tested impaired cell permeability directly or indirectly. Conclusions: 8PP and fluconazole and their combination caused alterations in cell permeability in azole-resistant C. albicans. They were effective both individually and in combination.

MgrB Mutations and Altered Cell Permeability in Colistin Resistance in Klebsiella pneumoniae.

There has been a resurgence in the clinical use of polymyxin antibiotics such as colistin due to the limited treatment options for infections caused by carbapenem-resistant Enterobacterales (CRE). However, this last-resort antibiotic is currently confronted with challenges which include the emergence of chromosomal and plasmid-borne colistin resistance. Colistin resistance in Klebsiella pneumoniae is commonly caused by the mutations in the chromosomal gene mgrB. MgrB spans the inner membrane and negatively regulates PhoP phosphorylation, which is essential for bacterial outer membrane lipid biosynthesis. The present review intends to draw attention to the role of mgrB chromosomal mutations in membrane permeability in K. pneumoniae that confer colistin resistance. With growing concern regarding the global emergence of colistin resistance, deciphering physical changes of the resistant membrane mediated by mgrB inactivation may provide new insights for the discovery of novel antimicrobials that are highly effective at membrane penetration, in addition to finding out how this can help in alleviating the resistance situation.

Mode of inactivation of Staphylococcus aureus and Escherichia coli by heated oyster-shell powder

• HOS is a highly effective disinfectant for the bacterial inactivation. • ROS species verification in HOS revealed the presence of singlet oxygen. • Disparate HOS disinfection rates between S. aureus and E. coli were firstly studied. • FM, TEM, and K + leakage provide early diagnosis of altered membrane permeability. • Biophysical properties and 3D images of inactivated bacteria were firstly identified. Bacterial infection and subsequent disinfection of microorganisms are ongoing issues around the world. Bio-calcium oxide derived from heated oyster-shell (HOS) waste product has been shown to be an effective disinfectant and has the additional advantage of the marketing use of utilizing waste materials that would otherwise contribute to environmental problems; however, its mode of inactivation is unknown. In this research, fluorescence microscopy (FM), transmission electron microscopy (TEM), atomic force microscopy (AFM), and electron spin resonance (ESR) spectrometer techniques were used to characterize the mode of inactivation of S. aureus and E. coli via HOS. This is the first work to provide insight into the three-dimensional morphology and biophysical properties of the inactivated S. aureus and E. coli cells via HOS. Noteworthy, the presence of singlet oxygen in HOS suspension altered bacterial cell permeability, leading to sustained inactivation. The HOS exhibited excellent disinfection capacity and achieved a 5-log-inactivation E. coli within 60 min with a dose of 0.2 g/L, superior to other shell-derived disinfectants. Thus, HOS provides a cost-effective disinfectant for the application of controlling pathogens.

Successful Management of an Aluminum Phosphide Poisoned Patient Following Ventricular Tachycardia

Aluminum Phosphide (ALP) poisoning is regarded as fatal; however, it is commonly used in developing countries. In some countries, such as India and Iran, it is still being used to protect grain against pests at home. As yet, no known antidote is reported to be effective against ALP poisoning. ALP acts through blockage of cell respiration, alteration of cell permeability, and induction of oxidative stress. Some factors can worsen the prognosis of its intoxication, which include hyperglycemia on admission, ingestion of fresh tablets, cardiac abnormality, especially Ventricular Tachycardia (VT), severe metabolic acidosis, cardiogenic shock and other types of shocks, electrolyte disturbances, end-organ damage, and a need for mechanical ventilation. ALP kills its victims by induction of multi-organ failure. In this study, we controlled the patient’s VT using MgSO4, electric shock, amiodarone, and lidocaine. We also encountered the cardiogenic shock using norepinephrine, High-Dose Insulin (HDI), antioxidants, sodium bicarbonate, and normal saline. Moreover, myocardial ischemia, recurrent VTs, and adverse cardiac effects were protected via tight control of magnesium and other electrolytes.

Anti-biofilm Potential and Mode of Action of Malaysian Plant Species: A Review

Biofilm is a microbial community that attaches to a surface and is enclosed in extracellular polymeric substance (EPS) matrix. Formation of biofilm often develops resistance towards a wide spectrum of antimicrobial agents. Since the biofilm-mediated diseases are commonly difficult to treat, there is a need to find new anti-biofilm agent. The studies on anti-biofilm activities of plant species have received a great deal of attention over the last few decades. In Malaysia, plant species have been used as alternatives to the conventional antimicrobial therapy. Several Malaysian plant species are known to control biofilm infection by inhibition of quorum sensing pathway, disruption of EPS matrix, alteration of cell permeability and reduction in cell surface hydrophobicity. This review demonstrates that Malaysian plant species may become excellent therapeutic agents in combating the biofilm infection.

Microbial esterases and ester prodrugs: An unlikely marriage for combating antibiotic resistance.

The rise of antibiotic resistance necessitates the search for new platforms for drug development. Prodrugs are common tools for overcoming drawbacks typically associated with drug formulation and delivery, with ester prodrugs providing a classic strategy for masking polar alcohol and carboxylic acid functionalities and improving cell permeability. Ester prodrugs are normally designed to have simple ester groups, as they are expected to be cleaved and reactivated by a wide spectrum of cellular esterases. However, a number of pathogenic and commensal microbial esterases have been found to possess significant substrate specificity and can play an unexpected role in drug metabolism. Ester protection can also introduce antimicrobial properties into previously nontoxic drugs through alterations in cell permeability or solubility. Finally, mutation to microbial esterases is a novel mechanism for the development of antibiotic resistance. In this review, we highlight the important pathogenic and xenobiotic functions of microbial esterases and discuss the development and application of ester prodrugs for targeting microbial infections and combating antibiotic resistance. Esterases are often overlooked as therapeutic targets. Yet, with the growing need to develop new antibiotics, a thorough understanding of the specificity and function of microbial esterases and their combined action with ester prodrug antibiotics will support the design of future therapeutics.

Hydroxypropyl-β-cyclodextrin-mediated alterations in cell permeability, lipid and protein profiles of steroid-transforming Arthrobacter simplex.

Hydroxypropyl-β-cyclodextrin (HP-β-CD) enhances steroid 1-dehydrogenation biotransformation by Arthrobacter simplex. In this work, HP-β-CD-induced improvement of A. simplex CPCC 140451 cell envelope permeability which had positive effects on the steroid bioconversion was confirmed by a comparative investigation which showed a lower dehydrogenase activity and higher cell permeability of the cells after being incubated with HP-β-CD. Atomic force microscopy and transmission electron microscopy micrographs showed that HP-β-CD altered the size, sharpness, and surface structure of the cell envelope. The analysis of lipid composition revealed that the proportion of extractable lipids decreased and the fatty acids profile was considerably altered. The contents of unsaturated fatty acids and long-chain fatty acids were reduced by 11.77 and 14.98%, respectively. The total leakage of protein level increased to 8%. Proteins belonging to the ATP-binding cassette superfamily and major facilitator superfamily were observed outside the cell. These alterations can explain the change of permeability on the molecular level under HP-β-CD treatment. Results showed the material basis and mechanisms underlying the cellular changes, thus most likely contributing to the conversion rate in addition to cyclodextrins known effects on substrate solubility.

Contribution of different mechanisms to the resistance to fluoroquinolones in clinical isolates of Salmonella enterica

ObjectivesTo study the potential factors include gene mutation, efflux pump and alteration of permeability associated with quinolone-resistance of Salmonella enterica strains isolated from patients with acute gastroenteritis and to evaluate the degree of synergistic activity of efflux pump inhibitors when combined with ciprofloxacin against resistant isolates. MethodsAntimicrobial resistance patterns of fifty-eight Salmonella isolates were tested. Five isolates were selected to study the mechanism of resistance associated with quinolone group, including mutation in topoisomerase-encoding gene, altered cell permeability, and expression of an active efflux system. In addition, the combination between antibiotics and efflux pump inhibitors to overcome the microbial resistance was evaluated. ResultsFive Salmonella isolates totally resistant to all quinolones were studied. All isolates showed alterations in outer membrane proteins including disappearance of some or all of these proteins (Omp-A, Omp-C, Omp-D and Omp-F). Minimum inhibitory concentration values of ciprofloxacin were determined in the presence/absence of the efflux pump inhibitors: carbonyl cyanide m-chlorophenylhydrazone, norepinephrin and trimethoprim. Minimum inhibitory concentration values for two of the isolates were 2–4 fold lower with the addition of efflux pump inhibitors. All five Salmonella isolates were amplified for gyrA and parC genes and only two isolates were sequenced. S. Enteritidis 22 had double mutations at codon 83 and 87 in addition to three mutations at parC at codons 67, 76 and 80 whereas S. Typhimurium 57 had three mutations at codons 83, 87 and 119, but no mutations at parC. ConclusionsEfflux pump inhibitors may inhibit the major AcrAB-TolC in Salmonella efflux systems which are the major efflux pumps responsible for multidrug resistance in Gram-negative clinical isolates.

Loss of surface antigens is a conserved feature of apoptotic lymphocytes from several mammalian species

Human lymphocytes lose the expression of lineage antigens (LAgs) along apoptosis. Our aim was to extent our previous studies of LAg loss to rodent species, quantifying LAg expression on apoptotic murine lymphocytes using flow cytometry to measure alterations in cell permeability, phosphatidylserine exposure and caspase activation of CD3, CD5, CD4, CD8, CD19 and CD28 LAgs in highly purified lymphocyte populations. We found loss of expression by apoptotic cells of all LAgs studied in the three species analyzed except for CD3 antigen in mouse. We also found an early, rapid and dramatic reduction in the expression of CD28 by early apoptotic cells. We found several homologies across the three species in the kinetic of loss of several LAgs such as CD5, CD4 and CD28. These data suggest that the loss of expression of LAgs by apoptotic lymphocytes is a common and conserved feature of lymphocytes undergoing apoptosis in several mammalian species.

Deletion of the histone-like protein (Hlp) from Mycobacterium smegmatis results in increased sensitivity to UV exposure, freezing and isoniazid

Adaptation to environmental stress is an important survival characteristic of any bacterial species. As a soil-dwelling saprophyte, Mycobacterium smegmatis is exposed to factors such as UV light and rounds of freezing and thawing that occur in temperate climates. Numerous studies in Escherichia coli have linked histone-like proteins to stress resistance and adaptation. We hypothesized that the 'histone-like' protein Hlp might likewise be involved in the stress response of M. smegmatis. The hlp gene was inactivated and the M. smegmatis Δhlp strain was found to be more susceptible to UV light and to the stress created by repeated cycles of freezing and thawing. In addition, loss of Hlp altered the colony morphology and allowed the organism to grow dispersed in the absence of a detergent, suggesting changes in the cell wall composition. As cell wall changes could affect permeability to certain antibiotics, the susceptibility of M. smegmatis Δhlp to kanamycin, rifamipicin, ethambutol and isoniazid (INH) was tested. M. smegmatis Δhlp was more susceptible to INH, but loss of Hlp did not affect susceptibility to the other antibiotics tested. This suggests that the increased sensitivity of M. smegmatis Δhlp to INH was unlikely to be the result of alterations in cell permeability.

BvrR/BvrS-Controlled Outer Membrane Proteins Omp3a and Omp3b Are Not Essential forBrucella abortusVirulence

The Brucella abortus two-component regulatory system BvrR/BvrS controls the expression of outer membrane proteins (Omp) Omp3a (Omp25) and Omp3b (Omp22). Disruption of bvrS or bvrR generates avirulent mutants with altered cell permeability, higher sensitivity to microbicidal peptides, and complement. Consequently, the role of Omp3a and Omp3b in virulence was examined. Similar to bvrS or bvrR mutants, omp3a and omp3b mutants displayed increased attachment to cells, indicating surface alterations. However, they showed unaltered permeability; normal expression of Omp10, Omp16, Omp19, Omp2b, and Omp1; native hapten polysaccharide; and lipopolysaccharide and were resistant to complement and polymyxin B at ranges similar to those of the wild-type (WT) counterpart. Likewise, omp3a and omp3b mutants were able to replicate in murine macrophages and in HeLa cells, were resistant to the killing action of human neutrophils, and persisted in mice, like the WT strain. Murine macrophages infected with the omp3a mutant generated slightly higher levels of tumor necrosis factor alpha than the WT, whereas the bvrS mutant induced lower levels of this cytokine. Since the absence of Omp3a or Omp3b does not result in attenuation, it can be concluded that BvrR/BvrS influences additional Brucella properties involved in virulence. Our results are discussed in the light of previous works suggesting that disruption of omp3a generates attenuated Brucella strains, and we speculate on the role of group 3 Omps.

Sjögren’s syndrome: a condition with features of chronic graft-versus-host disease: does duct cell adhesion or permeability play a role in pathogenesis?

Primary and Secondary Sjögren’s syndrome are disease complexes characterized by periductal inflammatory cell infiltration of the salivary and lacrimal glands and manifest as dry mouth and dry eyes. Secondary Sjögren’s syndrome may be associated with a connective tissue disorder. Additional extraglandular features in Sjögren’s syndrome include a generalized inflammatory exocrinopathy that might be associated with abnormalities of both humoral and cellular mediated immunity. Similar inflammatory changes and extraglandular features, including an altered immune response, have been reported in patients developing graft-versus-host disease after bone-marrow transplantation and in patients with primary biliary cirrhosis. The periductal nature of the inflammatory response involving minor salivary and other glands raises the possibility of altered duct cell adhesion or permeability in playing a role in the aetiopathogenesis of Sjögren’s syndrome. The paper pulls together evidence that could be interpreted in this light. Evidence for bacterial or viral factor(s) altering the antigenicity of the histocompartibility (HC) complex on ductal cells in Sjögren’s syndrome patients is also described. A hypothesis is proposed for Sjögren’s syndrome in which the principal feature is an alteration in salivary gland duct cell adhesion or permeability. A re-evaluation of current knowledge of these two conditions from a clinical and experimental context are interpreted in this light.

Anti-Candida and mode of action of two newly synthesized polymers: a modified poly (methylmethacrylate-co-vinylbenzoylchloride) and a modified linear poly (chloroethylvinylether-co-vinylbenzoylchloride) with special reference to Candida albicans and Candida tropicalis.

Polymeric antimicrobial agents represent a new and important direction that is developing in the field of antimicrobial agents. Antimicrobial activity of two newly synthesized polymers: a modified poly (methylmethacrylate-co-vinylbenzoylchloride) and a modified linear poly (chloroethylvinylether-co-vinylbenzoylchloride) have been investigated and found to be active. Both polymers have showed a broad antimicrobial activity against C. albicans and C. tropicalis. Minimal inhibitory concentrations (MIC's) for poly (methylmethacrylate-co-vinylbenzoyl chloride) were 100, 75 and 100 microg/ml in case of C. albicans (ATCC 2091), C. albicans (SC5314) and C. tropicalis, respectively. However, polycholoroethylvinylether-covinylbenzoylchloride inhibited C. albicans (ATCC 2091), C. albicans (SC5314) and C. tropicalis with minimum inhibitory concentration values (MIC's) of 150 microg/ml against the three tested Candida strains. Mode of action studies of both polymers on the medically important yeasts, C. albicans and C. tropicalis revealed that poly (methylmethacrylate-co-vinylbenzoylchloride) induced cytotoxicity, DNA damage, and altered cell permeability and morphology, which was manifested as aggregated and swollen yeast cells (C. albicans ATCC 2091) by fluorescent microscopy examination. Poly (chloroethylvinylether-co-vinylbenzoylchloride) increased cell permeability, and respiration for C. albicans and C. tropicalis. The tested polymers at 50 microg/ml had pronounced effects on C. albicans and C. tropicalis cell wall phosphopeptidomannane, proteins, sugars and phosphorus. Generally, the two polymers proved effective against the tested microorganisms, but growth inhibitory effect varied according to the composition of the polymer active group. Many investigators consider polymeric antimicrobial agents as a potential new approach for enhancing the efficiency of some existing antimicrobial agents, including prolonged activity, reduce their toxicity, as well as reduce the environmental issues associated with product use.

Approaches to detecting false positives in yeast two-hybrid systems.

While many novel associations predicted by two-hybrid library screens reflect actual biological associations of two proteins in vivo, at times the functional co-relevance of two proteins scored as interacting in the two-hybrid system is unlikely. The reason for this positive score remains obscure, which leads to designating such clones as false positives. After investigating the effect of over-expressing a series of putative false positives in yeast, we determined that expression of some of these clones induces an array of biological effects in yeast, including altered growth rate and cell permeability, that bias perceived activity of LacZ reporters. Based on these observations, we identify four simple strategies that can assist in determining whether a protein is likely to have been selected in a two-hybrid screen because of indirect metabolic effects.

Aetiology of oral cancer: alcohol

The effect of alcohol alone on the oral mucosa and its association with the development of oral cancer is difficult to establish, principally because alcohol consumption histories are difficult to verify, alter over time, both with respect to beverage type and quantity, and are frequently confounded by tobacco use. This review considers the various pathways by which alcohol may exert such an influence. Namely, due to topical exposure (e.g. direct effect on cell membranes, altered cell permeability, variation in enzymes that metabolise alcohol) and/or systemic effects (e.g. nutritional deficiency, immunological deficiency, disturbed liver function). Finally, the numerous papers that have sought to establish the relative risk for oral cancer in association with alcohol intake are reviewed.

The global problem of antimicrobial resistance.

Along with other multidrug-resistant pathogens previously identified, Streptococcus pneumoniae is becoming a serious concern in hospital and ambulatory care settings. The AIDS pandemic has increased the threat of resistance in Mycobacterium tuberculosis. Antimicrobial drug resistance involves three molecular mechanisms drug inactivation, altered cell permeability, and alteration of target sites. Surveillance and multidisciplinary approaches will be key to containing the threat of global antimicrobial resistance.

Increased chemocytotoxicity to colon cancer cells by shock wave-induced cavitation

Background/Aims: Cavitation has been shown to hinder colon cancer cell proliferation in vitro. This study aimed at investigating the interest of combining cavitation and cytotoxic drugs in vitro. Methods: HT-29 cells were exposed in suspension to cavitation (shock waves plus bubbles) before 5-fluorouracil (FUra) administration. Cytotoxicity was studied by means of clonogenic survival, cell proliferation by [3H]deoxyuridine ([3H] dUdR) incorporation, and influence of the treatments on the cell cycle by cytofluorimetry; the effects of cavitation on RNA incorporation of FUra, cell permeability, and activity of thymidilate synthetase (TS) were also studied. Results: A preliminary exposure to cavitation (as compared with FUra alone) induced decreased colony formation (by up to 2 log in certain conditions) and colony size. Cavitation alone induced increased incorporation of [3H]dUdR during 48 hours and stimulated TS activity, but in the presence of FUra, the concentration of the drug that causes 50% inhibition of control cell growth for [3H]dUdR incorporation was reduced by up to 1 log, and TS inhibition was increased after cavitation as compared with FUra alone. RNA incorporation of [14C]FUra was increased by cavitation, as a consequence of altered cell permeability rather than a direct RNA effect. Seventy-two hours after treatment, cavitation plus FUra decreased by more than 50% the S-phase fraction and also inhibited mitosis. Conclusions: Submitting HT-29 cells to cavitation before treatment by FUra significantly increases the effects of the drug. The action of both agents appears to be partially synergistic with a cycle specificity.

Downy Mildew of Pearl Millet: II. Extraction of Toxins and Alterations in Cell Permeability

Summary Phytopathogenic toxins produced after infection with Sclerospora graminicola , an obligate parasite, in pearl millet plants during pathogenesis were successfully extracted and partially purified with acetone. The acetone fractionated supernatant solution of green ear disease extracts contained high toxic metabolites, which reproduced the initial downy mildew disease symptoms in bioassay tests. Further, these toxic extracts contained a high cellulase enzyme activity and caused an increased loss of electrolytes from pearl millet (BJ 104) leaf tissues.