Bacteriophage DNA induces an interrupted immune response during phage therapy in a chicken model.

One of the hopes for overcoming the antibiotic resistance crisis is the use of bacteriophages to combat bacterial infections, the so-called phage therapy. This therapeutic approach is generally believed to be safe for humans and animals as phages should infect only prokaryotic cells. Nevertheless, recent studies suggested that bacteriophages might be recognized by eukaryotic cells, inducing specific cellular responses. Here we show that in chickens infected with Salmonella enterica and treated with a phage cocktail, bacteriophages are initially recognized by animal cells as viruses, however, the cGAS-STING pathway (one of two major pathways of the innate antiviral response) is blocked at the stage of the IRF3 transcription factor phosphorylation. This inhibition is due to the inability of RNA polymerase III to recognize phage DNA and to produce dsRNA molecules which are necessary to stimulate a large protein complex indispensable for IRF3 phosphorylation, indicating the mechanism of the antiviral response impairment.

Cellular Organelle-Related Transcriptomic Profile Abnormalities in Neuronopathic Types of Mucopolysaccharidosis: A Comparison with Other Neurodegenerative Diseases.

Mucopolysaccharidoses (MPS) are a group of diseases caused by mutations in genes encoding lysosomal enzymes that catalyze reactions of glycosaminoglycan (GAG) degradation. As a result, GAGs accumulate in lysosomes, impairing the proper functioning of entire cells and tissues. There are 14 types/subtypes of MPS, which are differentiated by the kind(s) of accumulated GAG(s) and the type of a non-functional lysosomal enzyme. Some of these types (severe forms of MPS types I and II, MPS III, and MPS VII) are characterized by extensive central nervous system disorders. The aim of this work was to identify, using transcriptomic methods, organelle-related genes whose expression levels are changed in neuronopathic types of MPS compared to healthy cells while remaining unchanged in non-neuronopathic types of MPS. The study was conducted with fibroblast lines derived from patients with neuronopathic and non-neuronopathic types of MPS and control (healthy) fibroblasts. Transcriptomic analysis has identified genes related to cellular organelles whose expression is altered. Then, using fluorescence and electron microscopy, we assessed the morphology of selected structures. Our analyses indicated that the genes whose expression is affected in neuronopathic MPS are often associated with the structures or functions of the cell nucleus, endoplasmic reticulum, or Golgi apparatus. Electron microscopic studies confirmed disruptions in the structures of these organelles. Special attention was paid to up-regulated genes, such as PDIA3 and MFGE8, and down-regulated genes, such as ARL6IP6, ABHD5, PDE4DIP, YIPF5, and CLDN11. Of particular interest is also the GM130 (GOLGA2) gene, which encodes golgin A2, which revealed an increased expression in neuronopathic MPS types. We propose to consider the levels of mRNAs of these genes as candidates for biomarkers of neurodegeneration in MPS. These genes may also become potential targets for therapies under development for neurological disorders associated with MPS and candidates for markers of the effectiveness of these therapies. Although fibroblasts rather than nerve cells were used in this study, it is worth noting that potential genetic markers characteristic solely of neurons would be impractical in testing patients, contrary to somatic cells that can be relatively easily obtained from assessed persons.

Bacteriophages-Dangerous Viruses Acting Incognito or Underestimated Saviors in the Fight against Bacteria?

The steadily increasing number of drug-resistant bacterial species has prompted the search for alternative treatments, resulting in a growing interest in bacteriophages. Although they are viruses infecting bacterial cells, bacteriophages are an extremely important part of the human microbiota. By interacting with eukaryotic cells, they are able to modulate the functioning of many systems, including the immune and nervous systems, affecting not only the homeostasis of the organism, but potentially also the regulation of pathological processes. Therefore, the aim of this review is to answer the questions of (i) how animal/human immune systems respond to bacteriophages under physiological conditions and under conditions of reduced immunity, especially during bacterial infection; (ii) whether bacteriophages can induce negative changes in brain functioning after crossing the blood-brain barrier, which could result in various disorders or in an increase in the risk of neurodegenerative diseases; and (iii) how bacteriophages can modify gut microbiota. The crucial dilemma is whether administration of bacteriophages is always beneficial or rather if it may involve any risks.

Efficacy of a Combination Therapy with Laronidase and Genistein in Treating Mucopolysaccharidosis Type I in a Mouse Model.

Mucopolysaccharidosis type I (MPS I) is a lysosomal storage disorder caused by α-L-iduronidase deficiency. The standard treatment, enzyme replacement therapy with laronidase, has limited effectiveness in treating neurological symptoms due to poor blood-brain barrier penetration. An alternative is substrate reduction therapy using molecules, such as genistein, which crosses this barrier. This study evaluated the effectiveness of a combination of laronidase and genistein in a mouse model of MPS I. Over 12 weeks, MPS I and wild-type mice received laronidase, genistein, or both. Glycosaminoglycan (GAG) storage in visceral organs and the brain, its excretion in urine, and the serum level of the heparin cofactor II-thrombin (HCII-T) complex, along with behavior, were assessed. The combination therapy resulted in reduced GAG storage in the heart and liver, whereas genistein alone reduced the brain GAG storage. Laronidase and combination therapy decreased liver and spleen weights and significantly reduced GAG excretion in the urine. However, this therapy negated some laronidase benefits in the HCII-T levels. Importantly, the combination therapy improved the behavior of female mice with MPS I. These findings offer valuable insights for future research to optimize MPS I treatments.

Induction of the mitochondrial pathway of apoptosis by enrofloxacin in the context of the safety issue of its use in poultry.

The overuse of antibiotics in both humans and livestock has led to the antibiotic resistance phenomenon which is now considered one of the biggest problems in the modern world. Some antibiotics used to control or prevent infections in livestock poultry were registered a long time ago, and as a result, data on the possible side effects of their use, both for birds and humans, are incomplete and should be updated. An example of such an antibiotic is enrofloxacin which has been widely used in poultry since 1989. Data in recent years have begun to indicate that this antibiotic induces the process of apoptosis in diverse types of eukaryotic cells. Unfortunately, such studies have never been conducted on chicken models even though it is in poultry that this antibiotic is most commonly used. Therefore, the purpose of this work was to investigate whether enrofloxacin induces apoptosis in chicken cells of the UMNSAH/DF-1 line and to study the molecular mechanism of its action. The results of these experiments indicated that enrofloxacin induces apoptosis in chicken cells but not in human HEK-293 and PC3 cells. This induction was accompanied by changes in the morphology and size of mitochondria, the process of apoptosome formation and activation of executive caspases, which clearly indicates the role of the mitochondrial pathway in the induction of apoptosis by enrofloxacin. This study is the first to show the toxicity of enrofloxacin against chicken cells and to demonstrate the exact mechanism of its action. The results presented in this work show the need to monitor the concentration of antibiotic residues in poultry foods as well as to study their impact on public health to guarantee consumer safety and prevent the phenomenon of antibiotic resistance in bacteria.

Analysis of Phage Regulatory RNAs: Sequencing Library Construction from the Fraction of Small Prokaryotic RNAs Less Than 50 Nucleotides in Length.

So far, bacterial regulatory sRNAs of length less than 50 nucleotides have been poorly understood, and a low number of such molecules has been identified. The first microRNA-size functional ribonucleic acid occurring in a bacterial cell has been described only recently, and it was found to be encoded by a bacteriophage. One of the reasons for such a scarcity in this field is the lack of procedures intended for the isolation and selection of molecules of this size from bacterial cells. To meet these difficulties, we describe here the few-step procedure of isolation, purification, selection, and sequencing library preparation that is dedicated to the fraction of very small, bacterial RNA molecules.

Complex effects of the exo-xis region of the Shiga toxin-converting bacteriophage Φ24B genome on the phage development and the Escherichia coli host physiology.

Lambdoid bacteriophages are excellent models in studies on molecular aspects of virus-host interactions. However, some of them carry genes encoding toxins which are responsible for virulence of pathogenic strains of bacteria. Shiga toxin-converting bacteriophages (Stx phages) encode Shiga toxins that cause virulence of enterohemorrhagic Escherichia coli (EHEC), and their effective production depends on Stx prophage induction. The exo-xis region of the lambdoid phage genome consists of genes which are dispensable for the phage multiplication under laboratory conditions; however, they might modulate the virus development. Nevertheless, their exact effects on the phage and host physiology remained unclear. Here, we present results of complex studies on the role of the exo-xis region of bacteriophage Φ24B, one of Stx2b phages. Transcriptomic analyses, together with proteomic and metabolomic studies, provided the basis for understanding the functions of the exo-xis region. Genes from this region promoted lytic development of the phage over lysogenization. Moreover, expression of the host genes coding for DnaK, DnaJ, GrpE, and GroELS chaperones was impaired in the cells infected with the Δexo-xis phage mutant, relative to the wild-type virus, corroborating the conclusion about lytic development promotion by the exo-xis region. Proteomic and metabolomic analyses indicated also modulation of gad and nrf operons, and levels of amino acids and acylcarnitines, respectively. In conclusion, the exo-xis region controls phage propagation and host metabolism by influencing expression of different phage and bacterial genes, directing the virus to the lytic rather than lysogenic developmental mode.

Bacteriophage-encoded 24B_1 molecule resembles herpesviral microRNAs and plays a crucial role in the development of both the virus and its host.

The 24B_1 small non-coding RNA molecule has been identified in Escherichia coli after induction of Shiga toxin-converting bacteriophage Φ24B. In this work, we focused on its direct role during phage and bacterial host development. We observed that in many aspects, this phage sRNA resembles herpesviral microRNAs. Similar to microRNAs, the mature 24B_1 is a short molecule, consisting of just 20 nucleotides. It is generated by cleaving the 80-nt long precursor transcript, and likely it undergoes a multi-step maturation process in which the Hfq protein plays an important role, as confirmed by demonstration of its binding to the 24B_1 precursor, but not to the 24B_1 mature form. Moreover, 24B_1 plays a significant role in maintaining the prophage state and reprogramming the host's energy metabolism. We proved that overproduction of this molecule causes the opposite physiological effects to the mutant devoid of the 24B_1 gene, and thus, favors the lysogenic pathway. Furthermore, the 24B_1 overrepresentation significantly increases the efficiency of expression of phage genes coding for proteins CI, CII, and CIII which are engaged in the maintenance of the prophage. It seems that through binding to mRNA of the sdhB gene, coding for the succinate dehydrogenase subunit, the 24B_1 alters the central carbon metabolism and causes a drop in the ATP intracellular level. Interestingly, a similar effect, called the Warburg switch, is caused by herpesviral microRNAs and it is observed in cancer cells. The advantage of the Warburg effect is still unclear, however, it was proposed that the metabolism of cancer cells, and all rapidly dividing cells, is adopted to convert nutrients such as glucose and glutamine faster and more efficiently into biomass. The availability of essential building blocks, such as nucleotides, amino acids, and lipids, is crucial for effective cell proliferation which in turn is essential for the prophage and its host to stay in the lysogenic state.

Correction of symptoms of Huntington disease by genistein through FOXO3-mediated autophagy stimulation.

Huntington disease (HD) is a neurodegenerative disorder caused by a mutation in the HTT gene. The expansion of CAG triplets leads to the appearance of misfolded HTT (huntingtin) forming aggregates and leading to impairment of neuronal functions. Here we demonstrate that stimulation of macroautophagy/autophagy by genistein (4',5,7-trihydroxyisoflavone or 5,7-dihydroxy-3-(4-hydroxyphenyl)-4 H-1-benzopyran-4-one) caused a reduction of levels of mutated HTT in brains of HD mice and correction of their behavior as assessed in a battery of cognitive, anxiety and motor tests, even if the compound was administered after symptoms had developed in the animals. Biochemical and immunological parameters were also improved in HD mice. Studies on molecular mechanisms of genistein-mediated stimulation of autophagy in HD cells indicated the involvement of the FOXO3-related pathway. In conclusion, treatment with genistein stimulates the autophagy process in the brains of HD mice, leading to correction of symptoms of HD, suggesting that it might be considered as a potential drug for this disease. Combined with a very recently published report indicating that impaired autophagy may be a major cause of neurodegenerative changes, these results may indicate the way to the development of effective therapeutic approaches for different neurodegenerative diseases by testing compounds (or possibly combinations of compounds) capable of stimulating autophagy and/or unblocking this process.Abbreviations: CNS: central nervous system; EPM: elevated plus-maze; GOT1/ASPAT: glutamic-oxaloacetic transaminase 1, soluble; GPT/ALAT/ALT: glutamic pyruvic transaminase, soluble; HD: Huntington disease; HTT: huntingtin; IL: interleukin; mHTT: mutant huntingtin; NOR: novel object recognition; MWM: Morris water maze; OF: open field; ROS: reactive oxygen species; TNF: tumor necrosis factor.

Therapeutic potential of lithium chloride and valproic acid against neuronopathic types of mucopolysaccharidoses through induction of the autophagy process.

Mucopolysaccharidoses (MPS) are a group of inherited disorders, caused by mutations in the genes coding for proteins involved (directly or indirectly) in glycosaminoglycan (GAG) degradation. A lack or drastically decreased residual activity of a GAG-degrading enzyme leads to the storage of these compounds, thus damaging proper functions of different cells, including neurons. The disease leads to serious psycho-motor dysfunctions and death before reaching the adulthood. Until now, induction of the autophagy process was considered as one of the therapeutic strategies for treatment of diseases caused by protein aggregation (Alzheimer's, Parkinson's, and Huntington's diseases). However, this strategy has only been recently suggested as a potential therapy for MPS. In this work, we show that the pharmacological stimulation of autophagy, by using valproic acid and lithium chloride, led to accelerated degradation of accumulated GAGs. Cytotoxicity tests indicated the safety of the use of the investigated compounds. We observed an increased number of lysosomes and enhanced degradation of heparan sulfate (one of GAGs). Induction of the autophagy process was confirmed by measuring abundance of the marker proteins, including LC3-II. Moreover, inhibition of this process resulted in abolition of the valproic acid- and LiCl-mediated reduction in GAG levels. This is the first report on the possibility of using valproic acid and lithium chloride for reducing levels of GAGs in neuronopathic forms of MPS.

Genomic Instability of G-Quadruplex Sequences in Escherichia coli: Roles of DinG, RecG, and RecQ Helicases.

Guanine-rich DNA can fold into highly stable four-stranded DNA structures called G-quadruplexes (G4). Originally identified in sequences from telomeres and oncogene promoters, they can alter DNA metabolism. Indeed, G4-forming sequences represent obstacles for the DNA polymerase, with important consequences for cell life as they may lead to genomic instability. To understand their role in bacterial genomic instability, different G-quadruplex-forming repeats were cloned into an Escherichia coli genetic system that reports frameshifts and complete or partial deletions of the repeat when the G-tract comprises either the leading or lagging template strand during replication. These repeats formed stable G-quadruplexes in single-stranded DNA but not naturally supercoiled double-stranded DNA. Nevertheless, transcription promoted G-quadruplex formation in the resulting R-loop for (G3T)4 and (G3T)8 repeats. Depending on genetic background and sequence propensity for structure formation, mutation rates varied by five orders of magnitude. Furthermore, while in vitro approaches have shown that bacterial helicases can resolve G4, it is still unclear whether G4 unwinding is important in vivo. Here, we show that a mutation in recG decreased mutation rates, while deficiencies in the structure-specific helicases DinG and RecQ increased mutation rates. These results suggest that G-quadruplex formation promotes genetic instability in bacteria and that helicases play an important role in controlling this process in vivo.

Enhanced Efficiency of the Basal and Induced Apoptosis Process in Mucopolysaccharidosis IVA and IVB Human Fibroblasts.

Morquio disease, also called mucopolysaccharidosis IV (MPS IV), belongs to the group of lysosomal storage diseases (LSD). Due to deficiencies in the activities of galactose-6-sulfate sulfatase (in type A) or ß-galactosidase (in type B), arising from mutations in GALNS or GLB1, respectively, keratan sulfate (one of glycosaminoglycans, GAGs) cannot be degraded efficiently and accumulates in lysosomes. This primary defect leads to many cellular dysfunctions which then cause specific disease symptoms. Recent works have indicated that different secondary effects of GAG accumulation might significantly contribute to the pathomechanisms of MPS. Apoptosis is among the cellular processes that were discovered to be affected in MPS cells on the basis of transcriptomic studies and some cell biology experiments. However, Morquio disease is the MPS type which is the least studied in light of apoptosis dysregulation, while RNA-seq analyses suggested considerable changes in the expression of genes involved in apoptosis in MPS IVA and IVB fibroblasts. Here we demonstrate that cytochrome c release from mitochondria is more efficient in MPS IVA and IVB fibroblasts relative to control cells, both under the standard cultivation conditions and after treatment with staurosporine, an apoptosis inducer. This indication of apoptosis stimulation was corroborated by measurements of the levels of caspases 9, 3, 6, and 7, as well as PARP, cleaved at specific sites, in Morquio disease and control fibroblasts. The more detailed analyses of the transcriptomic data revealed which genes related to apoptosis are down- and up-regulated in MPS IVA and IVB fibroblasts. We conclude that apoptosis is stimulated in Morquio disease under both standard cell culture conditions and after induction with staurosporine which may contribute to the pathomechanism of this disorder. Dysregulation of apoptosis in other MPS types is discussed.

Discriminating macromolecular interactions based on an impedimetric fingerprint supported by multivariate data analysis for rapid and label-free Escherichia coli recognition in human urine.

This manuscript presents a novel approach to address the challenges of electrode fouling and highly complex electrode nanoarchitecture, which are primary concerns for biosensors operating in real environments. The proposed approach utilizes multiparametric impedance discriminant analysis (MIDA) to obtain a fingerprint of the macromolecular interactions on flat glassy carbon surfaces, achieved through self-organized, drop-cast, receptor-functionalized Au nanocube (AuNC) patterns. Real-time monitoring is combined with singular value decomposition and partial least squares discriminant analysis, which enables selective identification of the analyte from raw impedance data, without the use of electric equivalent circuits. As a proof-of-concept, the authors demonstrate the ability to detect Escherichia coli in real human urine using an aptamer-based biosensor that targets RNA polymerase. This is significant, as uropathogenic E. coli is a difficult-to-treat pathogen that is responsible for the majority of hospital-acquired urinary tract infection cases. The proposed approach offers a limit of detection of 11.3 CFU/mL for the uropathogenic E. coli strain No. 57, an analytical range in all studied concentrations (up to 105 CFU/mL), without the use of antifouling strategies, yet not being specific vs other E.coli strain studied (BL21(DE3)). The MIDA approach allowed to identify negative overpotentials (-0.35 to -0.10 V vs Ag/AgCl) as most suitable for the analysis, offering over 80% sensitivity and accuracy, and the measurement was carried out in just 2 min. Moreover, this approach is scalable and can be applied to other biosensor platforms.

Clinical presentation of 13 children with alkaptonuria.

Until now, only a few studies have focused on the early onset of symptoms of alkaptonuria (AKU) in the pediatric population. This prospective, longitudinal study is a comprehensive approach to the assessment of children with recognized AKU during childhood. The study includes data from 32 visits of 13 patients (five males, eight females; age 4-17 years) with AKU. A clinical evaluation was performed with particular attention to eye, ear, and skin pigmentation, musculoskeletal complaints, magnetic resonance imaging (MRI), and ultrasound (US) imaging abnormalities. The cognitive functioning and adaptive abilities were examined. Molecular genetic analyses were performed. The most common symptoms observed were dark urine (13/13), followed by joint pain (6/13), and dark ear wax (6/13). In 4 of 13 patients the values obtained in the KOOS-child questionnaire were below the reference values. MRI and US did not show degenerative changes in knee cartilages. One child had nephrolithiasis. Almost half of the children with AKU (5/13) presented deficits in cognitive functioning and/or adaptive abilities. The most frequent HGD variants observed in the patients were c.481G>A (p.Gly161Arg) mutation and the c.240A>T (p.His80Gln) polymorphism. The newly described allele of the HGD gene (c.948G>T, p.Val316Phe) which is potentially pathogenic was identified.

Actin Cytoskeleton Polymerization and Focal Adhesion as Important Factors in the Pathomechanism and Potential Targets of Mucopolysaccharidosis Treatment.

The main approach used in the current therapy of mucopolysaccharidosis (MPS) is to reduce the levels of glycosaminoglycans (GAGs) in cells, the deposits considered to be the main cause of the disease. Previous studies have revealed significant differences in the expression of genes encoding proteins involved in many processes, like those related to actin filaments, in MPS cells. Since the regulation of actin filaments is essential for the intracellular transport of specific molecules, the process which may affect the course of MPSs, the aim of this study was to evaluate the changes that occur in the actin cytoskeleton and focal adhesion in cells derived from patients with this disease, as well as in the MPS I mouse model, and to assess whether they could be potential therapeutic targets for different MPS types. Western-blotting, flow cytometry and transcriptomic analyses were employed to address these issues. The levels of the key proteins involved in the studied processes, before and after specific treatment, were assessed. We have also analyzed transcripts whose levels were significantly altered in MPS cells. We identified genes whose expressions were changed in the majority of MPS types and those with particularly highly altered expression. For the first time, significant changes in the expression of genes involved in the actin cytoskeleton structure/functions were revealed which may be considered as an additional element in the pathogenesis of MPSs. Our results suggest the possibility of using the actin cytoskeleton as a potential target in therapeutic approaches for this disease.

Interactions and Insertion of Escherichia coli Hfq into Outer Membrane Vesicles as Revealed by Infrared and Orientated Circular Dichroism Spectroscopies.

The possible carrier role of Outer Membrane Vesicles (OMVs) for small regulatory noncoding RNAs (sRNAs) has recently been demonstrated. Nevertheless, to perform their function, these sRNAs usually need a protein cofactor called Hfq. In this work we show, by using a combination of infrared and circular dichroism spectroscopies, that Hfq, after interacting with the inner membrane, can be translocated into the periplasm, and then be exported in OMVs, with the possibility to be bound to sRNAs. Moreover, we provide evidence that Hfq interacts with and is inserted into OMV membranes, suggesting a role for this protein in the release of sRNA outside the vesicle. These findings provide clues to the mechanism of host-bacteria interactions which may not be defined solely by protein-protein and protein-outer membrane contacts, but also by the exchange of RNAs, and in particular sRNAs.

Were academic promotions in biochemistry and other research disciplines improperly controlled in Poland between 2011 and 2020? A response to the recently published "Who controls the national academic promotion system" article.

In the recently published article by Koza et al. (SAGE Open, 2023, 13, doi: 10.1177/21582440231177974), the authors analyzed the academic promotion system in Poland between 2011 and 2020. They concluded that "the Polish system of academic promotions in the past decade can hardly be regarded as based on pure merit", suggesting the impropriety, based on the participance of the members of the Central Board for Degrees and Titles in panels of experts evaluating the applications. Biochemistry was provided as a research discipline in which such an "impropriety" was the most pronounced, though other disciplines were only slightly less "improperly affected". Although the calculations presented by Koza and others (Koza et al., 2023) were proper, their conclusions were affected by fundamental errors in assessing the roles of the panelists and misinterpretation of the data. The drawbacks of the interpretations of the facts and in drawing conclusions are presented and discussed in this paper, underlining the necessity of being very careful when assessing any phenomenon and concluding about any mechanism. Indeed, only very well substantiated conclusions, strongly supported by objective data, should be published. This rule is very well known in biochemistry and other exact and natural sciences, and should be mandatory in all other research disciplines.

Sex-dependent differences in behavioral and immunological responses to antibiotic and bacteriophage administration in mice.

Introduction: The problem of antibiotic resistance is a global one, involving many industries and entailing huge financial outlays. Therefore, the search for alternative methods to combat drug-resistant bacteria has a priority status. Great potential is seen in bacteriophages which have the natural ability to kill bacterial cells. Bacteriophages also have several advantages over antibiotics. Firstly, they are considered ecologically safe (harmless to humans, plants and animals). Secondly, bacteriophages preparations are readily producible and easy to apply. However, before bacteriophages can be authorized for medical and veterinary use, they must be accurately characterized in vitro and in vivo to determinate safety. Methods: Therefore, the aim of this study was to verify for the first time the behavioral and immunological responses of both male and female mice (C57BL/6J) to bacteriophage cocktail, composed of two bacteriophages, and to two commonly used antibiotics, enrofloxacin and tetracycline. Animal behavior, the percentage of lymphocyte populations and subpopulations, cytokine concentrations, blood hematological parameters, gastrointestinal microbiome analysis and the size of internal organs, were evaluated. Results: Unexpectedly, we observed a sex-dependent, negative effect of antibiotic therapy, which not only involved the functioning of the immune system, but could also significantly impaired the activity of the central nervous system, as manifested by disruption of the behavioral pattern, especially exacerbated in females. In contrast to antibiotics, complex behavioral and immunological analyses confirmed the lack of adverse effects during the bacteriophage cocktail administration. Discussion: The mechanism of the differences between males and females in appearance of adverse effects, related to the behavioral and immune functions, in the response to antibiotic treatment remains to be elucidated. One might imagine that differences in hormones and/or different permeability of the blood-brain barrier can be important factors, however, extensive studies are required to find the real reason(s).

Applications of the phage display technology in molecular biology, biotechnology and medicine.

The phage display technology is based on the presentation of peptide sequences on the surface of virions of bacteriophages. Its development led to creation of sophisticated systems based on the possibility of the presentation of a huge variability of peptides, attached to one of proteins of bacteriophage capsids. The use of such systems allowed for achieving enormous advantages in the processes of selection of bioactive molecules. In fact, the phage display technology has been employed in numerous fields of biotechnology, as diverse as immunological and biomedical applications (in both diagnostics and therapy), the formation of novel materials, and many others. In this paper, contrary to many other review articles which were focussed on either specific display systems or the use of phage display in selected fields, we present a comprehensive overview of various possibilities of applications of this technology. We discuss an usefulness of the phage display technology in various fields of science, medicine and the broad sense of biotechnology. This overview indicates the spread and importance of applications of microbial systems (exemplified by the phage display technology), pointing to the possibility of developing such sophisticated tools when advanced molecular methods are used in microbiological studies, accompanied with understanding of details of structures and functions of microbial entities (bacteriophages in this case).

Contribution of vesicle trafficking dysregulation to the pathomechanism of mucopolysaccharidosis.

Although mucopolysaccharidoses (MPS) are monogenic diseases, caused by mutations in genes coding for enzymes involved in degradation of glycosaminoglycans (GAGs), recent studies suggested that changes in expressions of various genes might cause secondary and tertiary cellular dysfunctions modulating the course of these diseases. In this report, we demonstrate that vesicle trafficking regulation is affected in fibroblasts derived from patients suffering from 11 different types of MPS due to changes in levels of crucial proteins (estimated by automated Western-blotting) involved in this process, including caveolin, clathrin, huntingtin (Htt), APPL1, EEA1, GOPC, Rab5, and Rab7. Microscopic studies confirmed these results, while investigations of tissue samples derived from the MPS I mouse model indicated differences between various organs in this matter. Moreover, transcriptomic analyses provided a global picture for changes in expressions of genes related to vesicle trafficking in MPS cells. We conclude that vesicle trafficking is dysregulated in MPS cells and changes in this process might contribute to the molecular mechanisms of this disease. Most probably, primary GAG storage might cause a cellular stress response leading to dysregulation of expression of many genes which, in turn, results in changes in cellular processes like vesicle trafficking. This can significantly modulate the course of the disease due to enhancing accumulation of GAGs and altering crucial cellular processes. This hypothesis has been supported by normalization of levels of clathrin in MPS cells treated with either an active form of the deficient GAG-degrading enzyme or a compound (5,7-dihydroxy-3-(4-hydroxyphenyl)-4H-1-benzopyran-4-one) indirectly reducing the efficiency of GAG synthesis.