BackgroundHematological and biochemical abnormalities have been reported in patients with confirmed cases of coronavirus disease 2019 (COVID-19). This study compared selected hematological and biochemical parameters in patients with confirmed COVID-19 (RT-PCR positive) and suspected COVID-19 (RT-PCR negative).Patients and methodsA retrospective cross-sectional study was conducted among 106 patients admitted to the COVID-19 isolation centers at Ibn-Sina Hospital, Mukalla, Yemen, from 1 December 2021 to 30 October 2022. The study included 63 men and 43 women, with a mean age of 58.3 ± 17.5 years. Demographic, clinical, hematological, and biochemical data were extracted from medical records. The following were analyzed: complete blood count (CBC), random blood glucose (RBG), alanine transaminase (ALT), aspartate transaminase (AST), lactate dehydrogenase (LDH), gamma-glutamyl transferase (GGT), C-reactive protein (CRP), and ferritin.ResultsOf the 106 patients, 58 (54.72%) had confirmed (RT-PCR positive) COVID-19, and 48 (45.3%) had suspected (RT-PCR negative) COVID-19. Compared with the RT-PCR-negative group, the RT-PCR-positive patients had significantly higher hemoglobin, hematocrit, red blood cell (RBC), and white blood cell (WBC) counts and were more likely to exhibit neutrophilia, lymphopenia, and eosinopenia. Serum LDH, GGT, CRP, and ferritin levels were also significantly higher in the RT-PCR-positive group. Using binary logistic regression analysis, LDH (OR: 1.008, 95% CI: 1.002–1.013, P = 0.004) and GGT (OR: 0.949, 95% CI: 0.903–0.997, P = 0.039) were independently associated with RT-PCR positivity.ConclusionAmong the investigated laboratory markers, LDH and GGT showed independent associations with RT-PCR positivity. Given the retrospective cross-sectional design, these findings should be interpreted as associations rather than evidence of predictive or causal relationships.
Diabetes mellitus is a significant risk factor for urinary tract infections (UTIs), which occur more frequently, have a more severe course, and show a greater tendency to recur in affected individuals. Metabolic disturbances, particularly hyperglycemia and glucosuria, promote bacterial growth while simultaneously impairing innate and adaptive immune responses. In addition, diabetes-related complications, such as autonomic neuropathy, contribute to bladder dysfunction and urinary stasis, further facilitating bacterial colonization. Epidemiological evidence indicates a two- to four-fold increased risk of UTIs in patients with diabetes, along with a high prevalence of asymptomatic bacteriuria and recurrent infections. A key feature of this relationship is its bidirectional nature. UTIs can exacerbate glycemic dysregulation by triggering stress responses and increasing insulin resistance, leading to transient or sustained hyperglycemia. Uropathogenic Escherichia coli plays a critical role in the pathogenesis of UTIs, with hyperglycemic conditions further enhancing bacterial adhesion, biofilm formation, and immune evasion. Effective management requires simultaneous control of infection and optimization of glycemic control, particularly in the context of rising antimicrobial resistance. Emerging therapeutic strategies include targeting bacterial adhesion, disrupting biofilms, and modulating host immune responses. Improved understanding of host–pathogen interactions may enable the development of more effective preventive and therapeutic approaches in this high-risk population.
Connexin 36 (Cx36), a neuronal gap junction protein, is essential for retinal signal transmission but may also contribute to neurodegeneration by facilitating the spread of excitotoxic stress signals. This study aimed to examine the degradation pathway of Cx36 under stress conditions relevant to retinal disease. Using immortalized 661W murine photoreceptor cells transfected with GFP-tagged Cx36, we evaluated the effects of lysosomal inhibition (Bafilomycin A1) and excitotoxic stimulation (NMDA). Immunofluorescence revealed intracellular Cx36 accumulation following Bafilomycin and NMDA exposure, accompanied by increased expression of p62/SQSTM1 and PSMA1, suggesting impaired autophagic flux and a potential compensatory involvement of the proteasome. Elevated RAB11A levels suggested recruitment of endosomal recycling pathways. NMDA treatment was associated with increased ubiquitin accumulation, particularly in Cx36-overexpressing cells, indicating enhanced proteostatic stress under excitotoxic conditions. These findings highlight the stress-responsive behavior of Cx36 and suggest that connexin proteostasis may represent a potential target for future investigation in retinal neurodegeneration, including glaucoma and age-related macular degeneration. Immunofluorescence revealed intracellular accumulation of p62 and PSMA1 after Bafilomycin exposure with signal intensities increasing approximately 1.5-2-fold versus controls (n = 3, p < 0.05). These data are consistent with disrupted autophagic flux and suggest proteasome involvement, but do not constitute functional proof.
Polyacrylamide gel preparation for SDS–PAGE is conventionally initiated by the ammonium persulfate (APS)–TEMED system. Although effective, the relatively high volatility of TEMED may increase operator exposure during routine laboratory use. In this study, N,N,N′,N–tetraethylethylenediamine (TEEDA) was systematically evaluated as an alternative accelerator for SDS–PAGE gel preparation. TEEDA alone produced polymerization and protein separation performance comparable to those of the conventional TEMED–APS system, although higher concentrations (8.3 mM in 10% separating gels and 17.4 mM in 5% stacking gels) were required than those of TEMED (4.0 mM and 6.7 mM, respectively). When TEEDA concentrations comparable to those of TEMED were used, supplementation with 0.1% SBS in separating gels and 0.01% SBS in stacking gels restored polymerization and protein separation performance to levels comparable with those of the conventional TEMED–APS system. These findings provide an optimized TEEDA–based formulation for routine SDS–PAGE.
Objective:To characterize deoxyribonucleic acid (DNA), ribonucleic acid (RNA), and protein yields extracted from tissue of fourteen mouse organ types to establish baseline yield estimates for these biomolecules and provide guidance for tissue sample usage and storage in biomedical studies. Methods:Six wild-type C57BL/6J and six BALB/c nude mice were euthanized and fresh tissues were collected for extracting total DNA, RNA, and protein. The concentration, purity, and quality of nucleic acids were measured with NanoDrop™ One spectrophotometer, gel electrophoresis or capillary electrophoresis. Protein concentration was determined using the bicinchoninic acid (BCA) method. Results:DNA yields were highest in the colorectum and lowest in bone. RNA yields were highest in the colorectum and spleen of C57BL/6J and BALB/c nude mice, respectively, and lowest in bone. Protein yields were highest in the kidney and liver of C57BL/6J and BALB/c nude mice, respectively, and lowest in bone. The tissue quantities required to obtain 10 μg of DNA, 10 μg of RNA, and 1 mg of protein were calculated and presented along with corresponding tissue sizes. Conclusion:DNA, RNA, and protein yields varied significantly across different mouse tissue types. These findings provide a reference for estimating tissue sample sizes needed for downstream assays and optimizing the preservation of mouse tissues.
Mitochondrial dysfunction plays a critical role in the pathogenesis of cardiovascular and metabolic diseases. However, direct assessment of mitochondrial respiration in human vascular tissue remains technically challenging. In this study, we present an ex vivo approach for real-time analysis of mitochondrial respiration in human left internal mammary artery (LIMA) grafts obtained during coronary artery bypass grafting (CABG). LIMA segment was collected intraoperatively and processed for bioenergetic assessment using the Seahorse XF Flex 3D Analyzer. Mitochondrial respiration was evaluated using the Mito Stress Test, enabling real-time measurement of oxygen consumption rate (OCR). In parallel, tissue nucleotide levels were quantified using high-performance liquid chromatography (HPLC), allowing complementary assessment of cellular energy status and redox balance. We demonstrate the feasibility of measuring mitochondrial respiration in intact human arterial tissue ex vivo. To our knowledge, this is the first application of the Seahorse XF Flex 3D platform for real-time bioenergetic analysis in intact human vascular tissue. Combined analysis of OCR and nucleotide levels enabled integrated assessment of vascular bioenergetic status. The applied protocol enabled reliable assessment of key bioenergetic parameters, including basal and maximal respiration. This study establishes a novel proof-of-concept workflow for ex vivo bioenergetic profiling of human vascular grafts, providing a platform for future investigations of vascular metabolism in cardiovascular and metabolic disorders.
Metabolic dysfunction-associated kidney disease (MDAKD) is increasingly recognised as a major clinical consequence of the global rise in obesity, type 2 diabetes, hypertension, and cardiovascular disease. Accumulating evidence suggests that the gut microbiota may contribute to the development and progression of metabolic and renal disorders through complex mechanisms involving microbial metabolites, immune activation, and disruption of the intestinal barrier. This review provides an overview of current knowledge regarding the role of the gut microbiota and gut-derived metabolites in the pathogenesis of chronic kidney disease (CKD) associated with metabolic disorders. Special attention is given to short-chain fatty acids, bile acids, N-trimethylamine oxide, branched-chain amino acids, indoxyl sulfate, p-cresol sulfate, and lipopolysaccharides. Accumulating experimental and clinical evidence suggests that dysbiosis may contribute to chronic low-grade inflammation, insulin resistance, endothelial dysfunction, lipotoxicity, and profibrotic signaling pathways associated with kidney injury and cardiovascular complications. The review also identifies significant limitations in current microbiome research, such as the predominance of animal studies, methodological challenges in metabolite quantification, and difficulties in establishing causality in humans. Emerging therapeutic strategies targeting the gut microbiota, including dietary interventions, prebiotics, probiotics, sodium-glucose cotransporter 2 inhibitors, glucagon-like peptide-1 receptor agonists, and faecal microbiota transplantation, may offer novel approaches to slowing CKD progression and improving metabolic health. However, further mechanistic and clinical studies are required to determine the efficacy of microbiota-targeted therapies in MDAKD.
BackgroundUrine culture is the gold standard for diagnosing urinary tract infections (UTIs), but most samples yield negative results. A rapid and accurate screening method could accelerate reporting, reduce workload, and lower costs. This study aimed to evaluate the diagnostic performance of the MUS-3600 urine analyzer as a screening tool for UTIs.MethodsUrine samples submitted to the microbiology laboratory were analyzed using the MUS-3600 within 10 min of collection. The analyzer uses flow-type micro-imaging and artificial intelligence–based particle classification. Culture results with ≥104 CFU/mL were defined as positive; lower counts were negative.ResultsFor white blood cells (WBC, >39/μL), sensitivity, specificity, PPV, NPV, and elimination rate were 63.4%, 84.5%, 50.8%, 90.1%, and 67.5%, respectively. Bacilli (>103/μL) showed 62.1%, 88.1%, 56.9%, 90.2%, and 70.4%. Suspected cocci (>859/μL) demonstrated lower accuracy (44.4%, 78.7%, 34.5%, 84.9%, and 62.8%). Combining WBC and bacilli yielded the highest elimination rate (77.2%) and NPV (87.9%). The analyzer performed better in males (NPV 93.9%) than in females (NPV 82.1%).ConclusionThe MUS-3600 provides moderate accuracy for UTI screening and may help reduce unnecessary urine cultures, particularly in male patients. However, its limited sensitivity, especially for cocci, prevents its use as a stand-alone screening tool. Further studies are needed to confirm its role as a complementary method for culture elimination.
Candida glabrata (currently classified as Nakaseomyces glabratus) is an opportunistic fungal pathogen notable for its intrinsic antifungal tolerance and ability to persist in host environments. Although strain CBS138 has served as the principal model for genetic and functional studies, accumulating evidence indicates substantial intraspecies diversity that may shape virulence, immune interactions and stress adaptation. In particular, the widely used clinical isolate BG2 differs from CBS138 in genome structure, adhesin regulation and macrophage survival, yet the extent to which these differences are reflected at the fungal cell surface remains unknown. Here, we present a comparative characterization of the surface-exposed proteomes (surfaceomes) of CBS138 and BG2 across three biologically relevant growth conditions: YPD-grown yeast-like cells, RPMI-cultured planktonic aggregates and RPMI-formed biofilms. Using trypsin shaving combined with LC–MS/MS, we identified pronounced strain- and condition-dependent differences in surface protein composition, encompassing adhesins, yapsin proteases and selected moonlighting proteins. Whereas CBS138 showed greater representation of adhesion- and interaction-related surface proteins, BG2 preferentially displayed proteins associated with cell-wall architecture and remodelling, consistent with distinct surface-mediated adaptive strategies. Transmission electron microscopy revealed condition-dependent differences in cell-wall thickness in both strains, with BG2 displaying a broader range of values and the highest thickness under biofilm conditions, providing structural context for variation in protease accessibility and surface-protein detectability. Collectively, our findings highlight substantial surfaceome plasticity in C. glabrata and underscore the importance of considering intraspecies diversity when interpreting host–pathogen interactions and fungal virulence pathways.
Mucopolysaccharidosis (MPS) is a group of inherited metabolic diseases, characterized by defects in the degradation of glycosaminoglycans and their accumulation in lysosomes. However, various secondary cellular changes also contribute to the pathomechanism of MPS. Previous studies have reached contradictory conclusions about the changes in mitochondria in MPS, from increased numbers of mitochondria to impaired activities of some mitochondrial respiratory chain enzymes to no changes in mitochondrial respiration. In this preliminary, hypothesis-generating study, mitochondrial network morphology and mitochondrial DNA (mtDNA) abundance were investigated in fibroblasts derived from patients suffering from diverse MPS types. Fluorescence microscopy and real-time PCR were used to estimate these parameters, respectively. No significant changes in the mitochondrial network morphology were detected in MPS fibroblasts relative to control cells. Decreased levels of mtDNA relative to nuclear DNA levels were evident in some (I, II, IIIA, IIID, and VI) but not all MPS types compared to control fibroblasts. The results of this study suggest that there are some, although perhaps not dramatic, impairments of mitochondrial functions in some MPS types; however, they do not provide direct evidence of mitochondrial dysfunction. Therefore, these findings should be interpreted as descriptive and exploratory, highlighting the need for further functional and mechanistic studies.
Background:Tooth enamel and dentin are the most highly mineralized tissues in the human body, and their structural stability depends on the balanced content of calcium (Ca), phosphorus (P), and fluoride (F). Demineralization leads to the loss of these ions, weakening the tooth structure and increasing susceptibility to caries. Objective:This study aimed to evaluate elemental changes in Ca, P, and F in intact, demineralized, and remineralized teeth across the enamel, dentin-enamel junction and peripulpal dentin. Materials and methods:45 extracted human teeth were longitudinally sectioned to yield 90 paired specimens. One half of each tooth served as a baseline (intact) control, while the corresponding half was subjected to acid-induced demineralization. Following demineralization, the experimental halves were randomly assigned to one of three remineralization protocols (n = 15 per group): fluoride varnish, fluoride toothpaste with rinsing, and fluoride toothpaste without rinsing. Elemental composition was analyzed using scanning electron microscopy with energy-dispersive X-ray spectroscopy (SEM-EDX). Statistical analysis included paired t-tests for within-tooth comparisons and one-way ANOVA for treatment effects (p < 0.05). Results:At baseline, enamel exhibited significantly higher calcium and phosphorus levels compared to dentin regions. Demineralization resulted in a significant reduction in mineral content, particularly in dentin, along with complete loss of detectable fluoride (p < 0.0001). Remineralization increased calcium and fluoride levels across all groups. Fluoride varnish showed the highest fluoride retention trend, particularly in enamel; however, no statistically significant differences were observed among treatment groups for calcium and phosphorus. Conclusion:Fluoride-based treatments contributed to partial remineralization of tooth hard tissues. Fluoride varnish showed a tendency toward greater fluoride incorporation, particularly in enamel; however, statistically significant superiority over other treatment approaches was not demonstrated. These findings support the role of fluoride in remineralization while highlighting the need for further clinically relevant studies.
Mirror bacteria are organisms constructed from mirror-image biomolecules unlike those used by all known life and are moving from theoretical concept to practical feasibility. Their unusual chemistry that makes them resistant to natural degradation and isolated from ecosystems, fuels optimism for applications such as more stable medicines, long-term information storage, and novel platforms for bioengineering. At the same time, these very characteristics raise reservations: they challenge existing definitions of “life,” expose gaps in regulation, and risk undermining public trust if deployed without careful oversight. This perspective assesses not only those risks but also the opportunities of mirror bacteria, moving beyond narrow biosafety debates to explore their broader ethical, regulatory, and cultural implications. We argue that mirror bacteria represent a rare testcase for how science and society navigate the responsibilities of creating fundamentally new forms of life. Their development offers not only technical possibilities but also a chance to build more resilient frameworks for governance and ethics in synthetic biology.
Here, a simple and precise genome modification method for agrobacterium was described and tested in strain GV3101. Taking the induced endogenous Transfer DNA (T-DNA) as the single-strand homologous repair template, this makes it possible to achieve valid and accurate genome modification for agrobacterium even without the strengthening of gene editing tools such as the CRISPR/Cas9 system. In addition, the introduction of tool plasmids into recipient agrobacteria can be achieved by the freeze-thaw method rather than complex triparental mating or expensive electroporation, which makes it more convenient. Here, the modification rate of agrobacterium strain GV3101 at both the upstream and downstream of VirD2 is 4.5% with 100% sequence accuracy. This research provides a valuable potential tool for genetic engineering in agrobacterium, with potential applications in biotechnology and agricultural sciences.
The zebrafish (Danio rerio) has become a widely adopted vertebrate model in biomedical research, offering high translational value while supporting the principles of Replacement, Reduction, and Refinement (3Rs). Here, the Polish Zebrafish Society presents comprehensive, field-specific recommendations for the responsible and effective use of zebrafish across major areas of biomedical research. The document summarizes the scientific rationale and experimental advantages of zebrafish-based approaches in oncology, toxicology, neurology and neuropsychiatry, metabolic diseases, immunology, cardiology, and genetic disease modelling. The model’s key strengths include rapid development, optical transparency, genetic tractability, and strong conservation of molecular and physiological pathways relevant to human disease. These features enable real-time in vivo analysis of pathological processes, high-throughput pharmacological and toxicological screening, and functional validation of disease-associated genes. By integrating ethical considerations with robust experimental evidence, these recommendations aim to promote standardized implementation of zebrafish models in academic research, pharmaceutical development, and regulatory science. Broad adoption of zebrafish-based approaches can accelerate preclinical discovery, enhance translational relevance, and substantially reduce reliance on higher vertebrate models.
The role of proteins in cellular activity has received close attention. Proteome analysis can detect early enough changes in the cell before the appearance of pathological changes. Due to the increasing number of cardiovascular complications, more specifically hypertension in modern life, this experimental pilot study tries to detect early changes in the protein profile in cases with arterial hypertension. In this pilot study, animal models were used, specifically control rats and spontaneously hypertensive rats (SHRs), which are widely used as translational models relevant to human cardiovascular physiology. Using gel electrophoresis (GE) method, we analysed proteomic signature of blood plasma and aorta tissue samples of the animal models.
Extreme indoor and outdoor conditions in patients’ places of stay (hot or cold air and changes in humidity) lead to weakening of the lipid barrier function and disproportionate epidermal microbiota which results in a lower threshold of physical and chemical tolerance of the skin manifesting as erythema, burning, dryness and deterioration of skin condition. Chronic epidermal irritation caused by the above factors leads to discomfort and prevents the proper functioning of the skin, as it results in thickening of the epidermis, widening of the sebaceous glands and pigmentation disorders. Regenerative mechanisms preventing lipid peroxidation or carbonylation of skin cell proteins, including epidermis, are focused on restoring physiology and not on “fibroblast protection”, which can accelerate the loss of firmness. Taking into account the theory of free radical or telomeric aging, it may be stated that the skin exposed to extreme conditions ages faster. When faced with such a problem, it seems pointless to apply invasive rejuvenating treatments without prior diagnosis or acquiring basic knowledge of care and hygiene of the epidermis. Daily skin care and cosmetics play a key role in the anti-aging process. Hygiene and proper cosmetic habits are essential for health and youthful appearance of the skin. Appropriate regulatory exfoliation, or the so-called turnover of the epidermis, should be used so as not to generate excessive free radical stress. Antioxidants are necessary for this. Modern products based on a liquid crystal base compatible with the liquid crystal structure of epidermal cement together with active emollients allow for restoration of appropriate epidermis parameters. The method of exfoliation with selected ingredients enables long-term skin cleansing by accelerating the turnover of the epidermis without excessive free radical stress, drying and irritation.
Obesity, a global health catastrophe, arises from complex interactions between environmental factors and genetic predispositions. This review summarizes the current state of knowledge on the genetic basis of obesity and contrasts rare monogenic forms caused by mutations in a single gene with common polygenic forms caused by hundreds of genetic variants with small effects. We highlight important genes in neuroendocrine signaling pathways, particularly the leptin-melanocortin system involving MC4R, LEP, and POMC, as well as newly identified loci from genome-wide association studies such as FTO and SEC16B. The interplay between genetic probability and environmental factors underscores the heterogeneity of obesity phenotypes. Recent advances in pharmacotherapy, such as GLP-1 receptor agonists and dual/triple incretin agonists, demonstrate strong efficacy across various genetic backgrounds and underscore the translational relevance of genetic insights. New findings from different groups support the use of polygenic risk scores to identify individuals at risk and suggest prevention strategies. This review discusses the genomic data on clinical practice and emphasizes the possibilities and challenges of precision medicine in obesity treatment. Future research should focus on length of genetic screening and elucidating gene-environment interactions to optimize treatment outcomes.
Adeno-associated virus (AAV) vectors have become central to in vivo gene therapy across genetic and acquired diseases. Yet despite extensive preclinical validation, AAV programs often encounter translational gaps when advancing from rodents to non-human primates (NHPs) and ultimately to humans. These species barriers arise from differences in capsid-receptor interactions, intracellular trafficking, immune landscapes, and tissue microanatomy. Even within species, strain-level genetic variation can markedly alter vector performance, exemplified by the LY6A-dependent central nervous system (CNS) tropism of AAV-PhP.B in C57BL/6J mice. Old world monkeys, which are evolutionarily closer to humans than new world species, remain the most widely used models for systemic and CNS delivery, yet discrepancies in seroprevalence, complement activity, endothelial biology, and neuronal susceptibility still limit predictability. Recent advances, including machine learning-guided capsid design, deep mutational scanning, and human-derived organoids and explant models, offer powerful tools to bridge these barriers. This mini-review synthesizes current understanding of AAV species barriers and outlines strategies that enhance the robustness and human relevance of AAV gene therapy development.
ObjectiveBenign paroxysmal positional vertigo (BPPV) is a common vestibular disorder that responds well to canalith repositioning maneuvers but is frequently complicated by recurrence. Increasing evidence suggests that systemic inflammation may influence disease course and recurrence risk. This study aimed to investigate the association between routinely available systemic inflammatory markers and BPPV recurrence within 6 months.MethodsIn this single-center retrospective cohort study, 300 adult patients diagnosed with BPPV between January 2020 and December 2024 were included. Neutrophil-to-lymphocyte ratio (NLR), systemic immune-inflammation index (SII), and C-reactive protein (CRP) levels measured at initial presentation were analyzed. Recurrence within 6 months following successful canalith repositioning maneuvers was defined as the primary outcome. Group comparisons were performed using non-parametric tests. Independent predictors of recurrence were identified using multivariable logistic regression analysis, and receiver operating characteristic (ROC) analysis was used to evaluate discriminatory performance.ResultsDuring the six-month follow-up period, recurrence occurred in 78 patients (26.0%). Patients with recurrence had significantly higher NLR and CRP levels compared with those without recurrence (both p < 0.001). SII values were also elevated in the recurrence group, although the difference was less pronounced (p = 0.048). In multivariable analysis adjusted for age, sex, and canal involvement, both NLR (odds ratio [OR]: 1.42; 95% confidence interval [CI]: 1.18–1.71) and CRP (OR: 1.27; 95% CI: 1.10–1.46) emerged as independent predictors of BPPV recurrence. ROC analysis demonstrated moderate discriminatory ability for NLR (AUC = 0.71) and CRP (AUC = 0.69), whereas SII showed limited predictive performance (AUC = 0.62).ConclusionSystemic inflammatory markers, particularly NLR and CRP, are independently associated with recurrence risk in BPPV. Given their low cost and widespread availability, these markers may provide complementary information for early risk stratification. Further prospective and multicenter studies are required to clarify causal mechanisms and confirm clinical applicability.
