Lactoferricin B Combined with Antibiotics Exhibits Leukemic Selectivity and Antimicrobial Activity.

The fusion of penetrating peptides (PPs), e.g., cell penetration peptides (CPPs) or antimicrobial peptides (AMPs), together with antimicrobial agents is an expanding research field. Specific AMPs, such as lactoferricin B (LfcinB), have demonstrated strong antibacterial, antifungal, and antiparasitic activity, as well as valuable anticancer activity, proving beneficial in the development of anticancer conjugates. The resulting conjugates offer potential dual functionality, acting as both an anticancer and an antimicrobial agent. This is especially necessary in cancer treatment, where microbial infections pose a critical risk. Leukemic cells frequently exhibit altered outer lipid membranes compared to healthy cells, making them more sensitive to compounds that interfere with their membrane. In this study, we revisited and reanalyzed our earlier research on LfcinB and its conjugates. Furthermore, we carried out new experiments with a specific focus on cell proliferation, changes in membrane asymmetric phosphatidylserine location, intracellular reactive oxygen species (ROS) generation, mitochondrial functions, and in vitro bacterial topoisomerase inhibition.

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.

Personalized prediction of the secondary oocytes number after ovarian stimulation: A machine learning model based on clinical and genetic data.

Controlled ovarian stimulation is tailored to the patient based on clinical parameters but estimating the number of retrieved metaphase II (MII) oocytes is a challenge. Here, we have developed a model that takes advantage of the patient's genetic and clinical characteristics simultaneously for predicting the stimulation outcome. Sequence variants in reproduction-related genes identified by next-generation sequencing were matched to groups of various MII oocyte counts using ranking, correspondence analysis, and self-organizing map methods. The gradient boosting machine technique was used to train models on a clinical dataset of 8,574 or a clinical-genetic dataset of 516 ovarian stimulations. The clinical-genetic model predicted the number of MII oocytes better than that based on clinical data. Anti-Müllerian hormone level and antral follicle count were the two most important predictors while a genetic feature consisting of sequence variants in the GDF9, LHCGR, FSHB, ESR1, and ESR2 genes was the third. The combined contribution of genetic features important for the prediction was over one-third of that revealed for anti-Müllerian hormone. Predictions of our clinical-genetic model accurately matched individuals' actual outcomes preventing over- or underestimation. The genetic data upgrades the personalized prediction of ovarian stimulation outcomes, thus improving the in vitro fertilization procedure.

X-chromosome inactivation patterns depend on age and tissue but not conception method in humans.

Female somatic X-chromosome inactivation (XCI) balances the X-linked transcriptional dosages between the sexes, randomly silencing the maternal or paternal X chromosome in each cell of 46,XX females. Skewed XCI toward one parental X has been observed in association with ageing and in some female carriers of X-linked diseases. To address the problem of non-random XCI, we quantified the XCI skew in different biological samples of naturally conceived females of different age groups and girls conceived after in vitro fertilization (IVF). Generally, XCI skew differed between saliva, blood, and buccal swabs, while saliva and blood had the most similar XCI patterns in individual females. XCI skew increased with age in saliva, but not in other tissues. We showed no significant differences in the XCI patterns in tissues of naturally conceived and IVF females. The gene expression profile of the placenta and umbilical cord blood was determined depending on the XCI pattern. The increased XCI skewing in the placental tissue was associated with the differential expression of several genes out of 40 considered herein. Notably, skewed XCI patterns (> 80:20) were identified with significantly increased expression levels of four genes: CD44, KDM6A, PHLDA2, and ZRSR2. The differences in gene expression patterns between samples with random and non-random XCI may shed new light on factors contributing to the XCI pattern outcome and indicate new paths in future research on the phenomenon of XCI skewing.

Cellular and Gene Expression Response to the Combination of Genistein and Kaempferol in the Treatment of Mucopolysaccharidosis Type I.

Flavonoids are investigated as therapeutics for mucopolysaccharidosis, a metabolic disorder with impaired glycosaminoglycan degradation. Here we determined the effects of genistein and kaempferol, used alone or in combination, on cellular response and gene expression in a mucopolysaccharidosis type I model. We assessed the cell cycle, viability, proliferation, subcellular localization of the translocation factor EB (TFEB), number and distribution of lysosomes, and glycosaminoglycan synthesis after exposure to flavonoids. Global gene expression was analysed using DNA microarray and quantitative PCR. The type and degree of flavonoid interaction were determined based on the combination and dose reduction indexes. The combination of both flavonoids synergistically inhibits glycosaminoglycan synthesis, modulates TFEB localization, lysosomal number, and distribution. Genistein and kaempferol in a 1:1 ratio regulate the expression of 52% of glycosaminoglycan metabolism genes. Flavonoids show synergy, additivity, or slight antagonism in all analysed parameters, and the type of interaction depends on the concentration and component ratios. With the simultaneous use of genistein and kaempferol in a ratio of 4:1, even a 10-fold reduction in the concentration of kaempferol is possible. Flavonoid mixtures, used as the treatment of mucopolysaccharidosis, are effective in reducing glycosaminoglycan production and storage and show a slight cytotoxic effect compared to single-flavonoid usage.

Murine Fibroblasts and Primary Hepatocytes as Tools When Studying the Efficacy of Potential Therapies for Mucopolysaccharidosis Type I.

Mucopolysaccharidosis type I (MPS I) is a metabolic genetic disease caused by the deficiency of a lysosomal enzyme involved in glycosaminoglycans (GAGs) degradation. MPS I cells have a constant level of GAG synthesis, but disturbed degradation means that GAGs accumulate progressively, impairing cell metabolism. GAG metabolism can be modulated by flavonoids, and these are being studied as therapeutics for MPS. We have optimised the protocol for obtaining fibroblasts and hepatocytes from the MPS I murine model and characterised the cells for their suitability as an in vitro model for testing compounds with therapeutic potential. Methods: Murine primary hepatocytes and fibroblasts were used as a cellular model to study the effect of genistein, biochanin A, and kaempferol on the modulation of the GAG synthesis process. Flavonoids were used individually as well as in two-component mixtures. There were no statistically significant differences in GAG synthesis levels from cell types obtained from either wild-type or MPS I mice. We also showed that MPS I fibroblasts and hepatocytes store GAGs, which makes them useful in vitro models for testing the effectiveness of substrate reduction therapies. Furthermore, tested flavonoids had a different impact on GAG synthesis depending on cell type and whether they were used alone or in a mixture. The tested flavonoids reduce GAG synthesis more effectively in fibroblasts than in hepatocytes, regardless of whether they are used individually or in a mixture. Flavonoids modulate the level of GAG synthesis differently depending on cell types, therefore in vitro experiments performed to assess the effectiveness of potential therapies for metabolic diseases should be carried out using more than one cell model, and only such an approach will allow for full answering scientific questions.

Impact of isoflavone genistein on psoriasis in in vivo and in vitro investigations.

Genistein is applied worldwide as an alternative medicament for psoriasis (Ps) because of its anti-inflammatory activity and perceived beneficial impact on the skin. Hereby, we report our in vivo and in vitro investigations to supplement scientific research in this area. The reduction of clinical and biochemical scores in mild to moderate Ps patients taking genistein, its safety, good tolerability with no serious adverse events or discontinuations of treatment, no dose-limiting toxicities, negligible changes in pharmacodynamic parameters and remarkable serum interleukin level alterations were documented in this study. A certain regression of the Ps phenotype was visible, based on photo-documented Ps lesion evaluation. Through in vitro experiments, we found that genistein reduced IL-17A and TNF-α induced MAPK, NF-κB, and PI3K activation in normal human epidermal keratinocytes. Moreover, at the mRNA level of genes associated with the early inflammatory response characteristic for Ps (CAMP, CCL20, DEFB4A, PIK3CA, S100A7, and S100A9) and key cellular signalling (MTORC1 and TFEB), we showed that this isoflavone attenuated the increased response of IL-17A- and TNF-α-related pathways. This allows us to conclude that genistein is a good candidate for Ps treatment, being attractive for co-pharmacotherapy with other drugs.

Dosage Compensation in Females with X-Linked Metabolic Disorders.

Through the use of new genomic and metabolomic technologies, our comprehension of the molecular and biochemical etiologies of genetic disorders is rapidly expanding, and so are insights into their varying phenotypes. Dosage compensation (lyonization) is an epigenetic mechanism that balances the expression of genes on heteromorphic sex chromosomes. Many studies in the literature have suggested a profound influence of this phenomenon on the manifestation of X-linked disorders in females. In this review, we summarize the clinical and genetic findings in female heterozygotic carriers of a pathogenic variant in one of ten selected X-linked genes whose defects result in metabolic disorders.

Global Changes of 5-mC/5h-mC Ratio and Methylation of Adiponectin and Leptin Gene in Placenta Depending on Mode of Delivery.

It was suggested that the epigenetic alterations of the placenta are associated with obesity, as well as the delivery mode. This study aimed to assess the effect of maternal outcome and delivery procedure on global placental DNA methylation status, as well as selected 5'-Cytosine-phosphate-Guanine-3' (CpG) sites in ADIPOQ and LEP genes. Global DNA methylation profile in the placenta was assessed using the 5-methylcytosine (5-mC) and 5-hydroxymethylcytosine (5-hmC) ratio evaluated with the ELISA, followed by target gene methylation patterns at selected gene regions which were determined using methylation-specific qPCR in 70 placentas from healthy, pregnant women with single pregnancy. We found no statistically significant differences in 5-mC/5-hmC ratio between intrapartum cesarean sections (CS) and vaginal deliveries (p = 0.214), as well as between elective cesarean sections and vaginal deliveries (p = 0.221). In intrapartum cesarean sections, the ADIPOQ demethylation index was significantly higher (the average: 1.75) compared to elective cesarean section (the average: 1.23, p = 0.010) and vaginal deliveries (the average: 1.23, p = 0.011). The LEP demethylation index did not significantly differ among elective CS, intrapartum CS, and vaginal delivery groups. The demethylation index of ADIPOQ correlated negatively with LEP in the placenta in the vaginal delivery group (r = -0.456, p = 0.017), but not with the global methylation. The methylation of a singular locus might be different depending on the mode of delivery and uterine contractions. Further studies should be conducted with locus-specific analysis of the whole genome to detect the methylation index of specific genes involved in metabolism.

Lipophagy and Lipolysis Status in Lipid Storage and Lipid Metabolism Diseases.

This review discusses how lipophagy and cytosolic lipolysis degrade cellular lipids, as well as how these pathway ys communicate, how they affect lipid metabolism and energy homeostasis in cells and how their dysfunction affects the pathogenesis of lipid storage and lipid metabolism diseases. Answers to these questions will likely uncover novel strategies for the treatment of aforementioned human diseases, but, above all, will avoid destructive effects of high concentrations of lipids-referred to as lipotoxicity-resulting in cellular dysfunction and cell death.

The role of genetic factors and monocyte-to-osteoclast differentiation in the pathogenesis of Charcot neuroarthropathy.

Charcot neuroarthropathy is a chronic, progressive condition of the skeletal system that affects some patients with diabetic neuropathy. It results in progressive destruction of bones of the foot and disorganisation of pedal joints and ligaments. Effective prevention and treatment for Charcot neuroarthropathy remain a challenge. Currently, there are no reliable repeatable markers to identify patients with diabetes who are at higher risk of developing Charcot neuroarthropathy. The pathogenesis underlying the development of Charcot neuroarthropathy also remains unclear. In this review, we provide an overview of the history, prevalence, symptoms, risk factors, diagnostics and treatment of Charcot neuroarthropathy. We also discuss the potential for OPG and RANKL gene variants to act as predictive markers for the development of Charcot neuroarthropathy. Finally, we summarise the latest research on the role of monocyte-to-osteoclast differentiation in the development of acute Charcot neuroarthropathy.

Lipids and Lipid Mediators Associated with the Risk and Pathology of Ischemic Stroke.

Stroke is a severe neurological disorder in humans that results from an interruption of the blood supply to the brain. Worldwide, stoke affects over 100 million people each year and is the second largest contributor to disability. Dyslipidemia is a modifiable risk factor for stroke that is associated with an increased risk of the disease. Traditional and non-traditional lipid measures are proposed as biomarkers for the better detection of subclinical disease. In the central nervous system, lipids and lipid mediators are essential to sustain the normal brain tissue structure and function. Pathways leading to post-stroke brain deterioration include the metabolism of polyunsaturated fatty acids. A variety of lipid mediators are generated from fatty acids and these molecules may have either neuroprotective or neurodegenerative effects on the post-stroke brain tissue; therefore, they largely contribute to the outcome and recovery from stroke. In this review, we provide an overview of serum lipids associated with the risk of ischemic stroke. We also discuss the role of lipid mediators, with particular emphasis on eicosanoids, in the pathology of ischemic stroke. Finally, we summarize the latest research on potential targets in lipid metabolic pathways for ischemic stroke treatment and on the development of new stroke risk biomarkers for use in clinical practice.

Correction: Bochenska, K. et al. Lysosome Alterations in the Human Epithelial Cell Line HaCaT and Skin Specimens: Relevance to Psoriasis. Int. J. Mol. Sci. 2019, 20, 2255.

The authors wish to make the following corrections to this paper [...].

Lysosome Alterations in the Human Epithelial Cell Line HaCaT and Skin Specimens: Relevance to Psoriasis.

Despite the constantly updated knowledge regarding the alterations occurring in the cells of patients with psoriasis, the status and the role of the lysosome, a control center of cell metabolism, remain to be elucidated. The architecture of the epidermis is largely regulated by the action of lysosomes, possibly activating signaling pathways in the cellular crosstalk of keratinocytes-epidermal cells-with infiltrating immune cells. Thus, in the present study, lysosome alterations were examined in vitro and in situ using a two-dimensional (2D) keratinocyte model of HaCaT cells with "psoriasis-like" inflammation and skin specimens, respectively. Specific fluorescence and immunohistochemical staining showed an augmented level of acidic organelles in response to keratinocyte activation (mimicking a psoriatic condition while maintaining the membrane integrity of these structures) as compared with the control, similar to that seen in skin samples taken from patients. Interestingly, patients with the most pronounced PASI (Psoriasis Area and Severity Index), BSA (Body Surface Area), and DLQI (Dermatology Life Quality Index) scores suffered a high incidence of positive lysosomal-associated membrane protein 1 (LAMP1) expression. Moreover, it was found that the gene deregulation pattern was comparable in lesioned (PP) and non-lesioned (PN) patient-derived skin tissue, which may indicate that these alterations occur prior to the onset of the characteristic phenotype of the disease. Changes in the activity of genes encoding the microphthalmia family (MiT family) of transcription factors and mammalian target of rapamycin complex 1 (MTORC1) were also observed in the in vitro psoriasis model, indicating that the biogenesis pathway of this arm is inhibited. Interestingly, in contrast to the keratinocytes of HaCaT with "psoriasis-like" inflammation, LAMP1 was up-regulated in both PP and PN skin, which can be a potential sign of an alternative mechanism of lysosome formation. Defining the molecular profile of psoriasis in the context of "the awesome lysosome" is not only interesting, but also desired; therefore, it is believed that this paper will serve to encourage other researchers to conduct further studies on this subject.

The Role of Dimethyl Sulfoxide (DMSO) in Gene Expression Modulation and Glycosaminoglycan Metabolism in Lysosomal Storage Disorders on an Example of Mucopolysaccharidosis.

Obstacles to effective therapies for mucopolysaccharidoses (MPSs) determine the need for continuous studies in order to enhance therapeutic strategies. Dimethyl sulfoxide (DMSO) is frequently utilised as a solvent in biological studies, and as a vehicle for drug therapy and the in vivo administration of water-insoluble substances. In the light of the uncertainty on the mechanisms of DMSO impact on metabolism of glycosaminoglycans (GAGs) pathologically accumulated in MPSs, in this work, we made an attempt to investigate and resolve the question of the nature of GAG level modulation by DMSO, the isoflavone genistein solvent employed previously by our group in MPS treatment. In this work, we first found the cytotoxic effect of DMSO on human fibroblasts at concentrations above 3%. Also, our results displayed the potential role of DMSO in the regulation of biological processes at the transcriptional level, then demonstrated a moderate impact of the solvent on GAG synthesis. Interestingly, alterations of lysosomal ultrastructure upon DMSO treatment were visible. As there is growing evidence in the literature that DMSO can affect cellular pathways leading to numerous changes, it is important to expand our knowledge concerning this issue.

Molecular action of isoflavone genistein in the human epithelial cell line HaCaT.

Due to its strong proliferation-reducing effects on keratinocytes, and also anti-inflammatory properties, the isoflavone genistein has already been proposed as a possible antipsoriatic compound. As there is still no detailed information on this topic, we examined the effects of genistein by using an in vitro model of both, normal and "psoriasis-like" keratinocytes at this stage of our work exhaustively testing the selected flavonoid in a mono-treated experimental design. Gene expression studies revealed transcriptional changes that confirms known disease-associated pathways and highlights many psoriasis-related genes. Our results suggested that aberrant expression of genes contributing to the progress of psoriasis could be improved by the action of genistein. Genistein prevented "cytokine mix" as well as TNF-α-induced NF-κB nuclear translocation, with no effect on the PI3K signaling cascade, indicating the luck of turning this pathway into NF-κB activation. It could have attenuated TNF-α and LPS-induced inflammatory responses by suppressing ROS activation. Regardless of the type of keratinocyte stimulation used, reduction of cytokine IL-8, IL-20 and CCL2 production (both at RNA and protein level) following genistein treatment was visible. Because investigations of other groups supported our commentary on potential administration of genistein as a potential weapon in the armamentarium against psoriasis, it is believed that this paper should serve to encourage researchers to conduct further studies on this subject.

Abnormal Sphingolipid World in Inflammation Specific for Lysosomal Storage Diseases and Skin Disorders.

Research in recent years has shown that sphingolipids are essential signalling molecules for the proper biological and structural functioning of cells. Long-term studies on the metabolism of sphingolipids have provided evidence for their role in the pathogenesis of a number of diseases. As many inflammatory diseases, such as lysosomal storage disorders and some dermatologic diseases, including psoriasis, atopic dermatitis and ichthyoses, are associated with the altered composition and metabolism of sphingolipids, more studies precisely determining the responsibilities of these compounds for disease states are required to develop novel pharmacological treatment opportunities. It is worth emphasizing that knowledge from the study of inflammatory metabolic diseases and especially the possibility of their treatment may lead to insight into related metabolic pathways, including those involved in the formation of the epidermal barrier and providing new approaches towards workable therapies.

Non-steroidal anti-inflammatory drugs are safe with respect to the transcriptome of human dermal fibroblasts.

Non-steroidal anti-inflammatory drugs (NSAIDs) provide important benefits to millions of patients, but are associated with a number of serious adverse events. These adverse drug reactions are an important clinical issue and a serious public health risk. While most unfortunate responses in human to NSAIDs are mild and may disappear after decreasing the dose or withdrawal of the drug, some of them can produce serious outcomes. Currently, little is known regarding the effects of NSAIDs on global RNA expression in normal, non-transformed cells. Therefore, in this report, the effect of NSAIDs, COX-nonspecific and COX-2-specific inhibitors, indomethacin and nimesulide respectively, commonly used medications worldwide for the reduction of pain, fever, inflammation and stiffness, on transcriptomic signature of human dermal fibroblasts was investigated. A total of 3803 differentially expressed genes with a fold change greater than or equal to 1.3 and below than or equal to 0.7 for whole genome transcripts, with a P value of < 0.05 were identified in response to all applied conditions. We found that although the total number of deregulated genes was relatively high at such criteria, changes in fibroblast transcriptome profile after treatment at selected experimental conditions were however smallish, as the selected drugs slightly modulate transcriptome with only a few genes with expression altered a bit more than twice. Nevertheless, transcriptomic data has its own limitations and it cannot reflect all post-transcriptional changes, which in turn may cause same risks, especially for a long time of medication.

Female Fabry disease patients and X-chromosome inactivation.

Fabry disease is an X-linked inherited lysosomal storage disorder caused by mutations in the gene encoding α-galactosidase A (GLA). Once it was thought to affect only hemizygous males. Over the last fifteen years, research has shown that most females carrying mutated allele also develop symptoms, demonstrating a wide range of disease severity, from a virtually asymptomatic to more classical profile, with cardiac, renal, and cerebrovascular manifestations. This variable expression in females is thought to be influenced by the process of X-chromosome inactivation (XCI). The aim of this study was to assess severity of the clinical phenotype, to analyze XCI patterns, and to estimate their effect on disease manifestation in twelve female Fabry disease patients from five unrelated Polish families. Our analyses revealed that patients presented with the broad range of disease expression - from mild to severe, and their clinical involvement did not correlate with XCI profiles. Female carriers of the mutation in the GLA gene with the random XCI may present with the wide range of disease signs and symptoms. Thus, XCI is not a main factor in the phenotype variability of Fabry disease manifestation in heterozygous females.

Models in the Research Process of Psoriasis.

Psoriasis is an ancient, universal chronic skin disease with a significant geographical variability, with the lowest incidence rate at the equator, increasing towards the poles. Insights into the mechanisms responsible for psoriasis have generated an increasing number of druggable targets and molecular drugs. The development of relevant in vitro and in vivo models of psoriasis is now a priority and an important step towards its cure. In this review, we summarize the current cellular and animal systems suited to the study of psoriasis. We discuss the strengths and limitations of the various models and the lessons learned. We conclude that, so far, there is no one model that can meet all of the research needs. Therefore, the choice model system will depend on the questions being addressed.