Search Results (159)

Lactic acid bacteria (LAB) are central to food, feed, and health biotechnology, yet their genomes have long resisted rapid, precise manipulation. This review charts the evolution of LAB genome-editing strategies from labor-intensive RecA-dependent double-crossovers to state-of-the-art CRISPR and CRISPR-associated transposase systems. Native homologous recombination, transposon mutagenesis, and phage-derived recombineering opened the door to targeted gene disruption, but low efficiencies and marker footprints limited throughput. Recent phage RecT/RecE-mediated recombineering and CRISPR/Cas counter-selection now enable scar-less point edits, seamless deletions, and multi-kilobase insertions at efficiencies approaching model organisms. Endogenous Cas9 systems, dCas-based CRISPR interference, and CRISPR-guided transposases further extend the toolbox, allowing multiplex knockouts, precise single-base mutations, conditional knockdowns, and payloads up to 10 kb. The remaining hurdles include strain-specific barriers, reliance on selection markers for large edits, and the limited host-range of recombinases. Nevertheless, convergence of phage enzymes, CRISPR counter-selection and high-throughput oligo recombineering is rapidly transforming LAB into versatile chassis for cell-factory and therapeutic applications. Full article

Arrhythmogenic cardiomyopathies (ACMs) are a phenotypically and etiologically heterogeneous group of myocardial disorders characterized by fibrotic or fibro-fatty replacement of ventricular myocardium, electrical instability, and an elevated risk of sudden cardiac death. Initially identified as a right ventricular disease, ACMs are now recognized to include biventricular and left-dominant forms. Genetic causes account for a substantial proportion of cases and include desmosomal variants, non-desmosomal variants, and familial gene-elusive forms with no identifiable pathogenic mutation. Nongenetic etiologies, including post-inflammatory, autoimmune, and infiltrative mechanisms, may mimic the phenotype. In many patients, the disease remains idiopathic despite comprehensive evaluation. Cardiac magnetic resonance imaging has emerged as a key tool for identifying non-ischemic scar patterns and for distinguishing arrhythmogenic phenotypes from other cardiomyopathies. Emerging classifications propose the unifying concept of scarring cardiomyopathies based on shared structural substrates, although global consensus is evolving. Risk stratification remains challenging, particularly in patients without overt systolic dysfunction or identifiable genetic markers. Advances in tissue phenotyping, multi-omics, and artificial intelligence hold promise for improved prognostic assessment and individualized therapy. Full article

Cardiac fibrosis following myocardial infarction plays a critical role in the formation of scar tissue and contributes to ventricular arrhythmias, including ventricular tachycardia and sudden cardiac death. Current clinical diagnostics use electrical and structural markers, but lack precision due to low spatial resolution and absence of molecular information. In this paper, we employed line scan Raman microspectroscopy to classify sheep myocardial tissue into muscle, necrotic, granulated, and fibrotic tissue types, using collagen as a molecular biomarker. Three spectral regions were evaluated: region A (600–2960 cm⁻¹), region B (600–1399 cm⁻¹ and 1751–2960 cm⁻¹), and region C (1400–1750 cm⁻¹), which includes the prominent collagen-associated peaks at 1448 cm⁻¹ and 1652 cm⁻¹. Linear and nonlinear principal component analysis (PCA) and support vector machines (SVMs) were applied for dimensionality reduction and classification, with nonlinear models specifically addressing the nonlinearity of collagen formation during fibrogenesis. Histological validation was performed using Masson’s trichrome staining. Raman bands associated with collagen in region C consistently outperformed regions A and B, achieving the highest explained variance and best class separation in both binary and multiclass PCA models for both linear and nonlinear approaches. The ratio of collagen-related peaks enabled stage-dependent tissue characterization, confirming the nonlinear nature of fibrotic remodeling. Our findings highlight the diagnostic potential of collagen-associated Raman bands for characterizing myocardial fibrosis. The proposed PCA-SVM framework demonstrates robust performance even with limited sample size and has the potential to lay the foundation for real-time intraoperative diagnostics. Full article

Intracerebral hemorrhage (ICH), a severe stroke subtype common in the elderly, often results in high morbidity and mortality, with limited treatment options for long-term recovery. While glial scar formation is increasingly recognized as key to central nervous system (CNS) repair, its role and characteristics in the aging brain post-ICH remain unclear. This study investigated glial scar formation after ICH (100 μL autologous blood injected into the right basal ganglia model) in aged Fischer 344 rats and assessed the effects of deferoxamine (DFX) treatment. Histological and immunohistochemical analyses were conducted on days 7, 28, and 60 post-ICH using cell-specific and iron-related markers, with DFX administered at 100 mg/kg daily for 14 days in separate groups. Over time, the lesion core showed increased hemosiderin accumulation and astrogliosis. By day 60, the area of astrogliosis corresponded to an area with persistent neuronal loss (DARPP-32-negative). Glial composition shifted from microglia dominance on day 28 to astrocyte predominance by day 60. DFX treatment reduced iron deposition, astrogliosis, and DARPP-32-negative regions while enhancing oligodendrocyte presence. Iron-related markers (HO-1, ferritin, Perls’ staining) and PDGFRβ-positive fibrotic cells were concentrated in the scar core. These findings provide novel insights into scar formation after ICH in aged rats and suggest DFX as a potential therapy to improve outcomes in elderly stroke patients. Full article

Root-knot nematodes, Meloidogyne spp., are widespread phytoparasites that cause a significant reduction in the yield of tomato Solanum lycopersicum. In the Russian Federation, where the use of chemical nematicides is limited due to environmental and toxicological risks, the cultivation of resistant varieties and hybrids remains the most effective and environmentally safe method to control Meloidogyne. In the course of this study, the resistance screening of 20 tomato varieties/hybrids and 21 mutant lines from the collection of the FSBSI FRCBPP to M. hapla was carried out using a comprehensive approach that included morphological and biochemical analysis methods. Resistance was assessed by calculating the gall formation index, the degree of root system damage, and biochemical parameters of fruits—vitamin C content and titratable acidity. In addition, molecular screening was carried out using the SCAR marker Mi23 to identify the Mi-1.2 gene, known as a key factor in resistance to a number of Meloidogyne spp. Although Mi-1.2 is not typically associated with resistance to M. hapla, all genotypes carrying this gene showed phenotypic resistance. This unexpected correlation suggests the possible involvement of Mi-associated or parallel mechanisms and highlights the need for further investigation into noncanonical resistance pathways. It was found that when susceptible genotypes were infected with M. hapla, there was a tendency for the vitamin C content to decrease, while resistant lines retained values close to the control. The presence of the Mi-1.2 gene was confirmed in 9.5% of samples. However, the phenotypic resistance of some lines, such as Volgogradets, which do not contain a marker for the Mi-1.2 gene, indicates a polygenic nature of resistance, alternative genetic mechanisms, or the possible influence of epigenetic mechanisms. The obtained data highlight the potential of using the identified resistant genotypes in breeding programs and the need for further studies of the molecular mechanisms of resistance, including the search for new markers specific to M. hapla, to develop effective strategies for tomato protection in sustainable agriculture. Full article

Background/Objectives: The aim of this study is to identify markers and develop a multifactorial model for characterizing extensive scar tissue after revascularization in patients with myocardial infarction (MI). Methods: A total of 123 patients with MI were examined. The patients underwent contrast-enhanced cardiac magnetic resonance imaging (MRI) with a 1.5 Tesla GE SIGNA Voyager (GE HealthCare, Chicago, IL, USA) on the 7th–10th days from the onset of the disease. At the first stage, we performed a comparative analysis and built a multifactorial model based on the examination results of 92 (75%) patients enrolled from April 2021 to October 2023. These patients formed the group used for model development, or the “modeling group”. The mass of the scar was calculated, including relative to the left ventricular (LV) myocardium mass (M_scar/LVMM, in %). Results: The first subgroup consisted of 36 (39%) patients with a large scar, denoted as “LS” (M_scar/LVMM > 20%). The second subgroup included 56 (61%) patients with a smaller scar, referred to as “SS” (M_scar/LVMM ≤ 20%). Logistic regression was used to identify independent factors affecting scar tissue size. A multifactorial model was created. This model predicts M_scar/LVMM > 20% on MRI. It uses readily available clinical parameters: high-sensitivity troponin I (HscTn I) and N-terminal pro B-type natriuretic peptide (NT-proBNP) levels, and LV relative wall thickness (RWT). We tested the multifactorial model on the “modeling group” (n = 31). The sensitivity was 63.6% and the specificity was 85.7%. Conclusions: These indicates the feasibility of its application in clinical practice. Full article

Peritoneal adhesions following surgical injury remain a major clinical challenge, often resulting in severe complications, such as intestinal obstruction, chronic pain, and infertility. This review systematically integrates recent genomic and molecular biology insights into the pathogenesis of peritoneal adhesions, explicitly focusing on molecular pathways, including TGF-β signaling, COX-2-mediated inflammatory responses, fibrinolytic balance (tPA/PAI-1), angiogenesis pathways (VEGF, PDGF), and extracellular matrix remodeling (MMPs/TIMPs). Newly conducted transcriptomic and proteomic analyses highlight distinct changes in gene expression patterns in peritoneal fibroblasts during adhesion formation, pinpointing critical roles for integrins, cadherins, selectins, and immunoglobulin superfamily molecules. Recent studies indicate significant shifts in TGF-β isoforms expression, emphasizing isoform-specific impacts on fibrosis and scarring. These insights reveal substantial knowledge gaps, particularly the differential regulatory mechanisms involved in fibrosis versus normal reparative reperitonealization. Future therapeutic strategies could target these molecular pathways and inflammatory mediators to prevent or reduce adhesion formation. Further research into precise genetic markers and the exploration of targeted pharmacological interventions remain pivotal next steps in mitigating postoperative adhesion formation and improving clinical outcomes. Full article

Background/Objectives: Saussurea obvallata (DC.) Edgew., Asteraceae, is a traditional medicinal herbnative to the Qinghai–Tibet Plateau (QTP). Pharmacological investigationshave validated its pharmacological effects in anti-tumor, anti-inflammatory, heat-clearing, detoxifying, and analgesia. S. obv is presently facing habitat fragmentation and population decline. Therefore, we analyzed its genetic and chemical diversity to provide a scientific basis for the conservation and sustainable use of S. obv. Methods: Seven populations of S. obv were sampled from Xizang, China. The genetic diversity was analyzed using inter-simple sequence repeat (ISSR) markers, and metabolites were identified by ultra-high-performance liquid chromatography-tandem-quadrupole-time-of-flight mass spectrometry (UPLC-Q-TOF-MS/MS). Correlation analysis among genetic diversity, differential metabolites, and climatic factors were performed by R. Results: The genetic diversity among and within populations were both lowly and significantly correlated with geographical distance, showing a decreasing trend from east to west of the QTP. A total of 110 compounds were identified, including flavonoids, phenylpropanoids, lipids, fatty acids, terpenoids, alkaloids, etc. The metabolite contents among populations varied greatly and were related to environmental factors, mainly annual mean temperature and temperature fluctuation. The genetic diversity had little effect on the metabolic differences. Conclusions: These findings provided valuable baseline information for the conservation and pharmacological utilization of S. obv. Meanwhile, further research is necessary for the efficacy evaluation of anti-inflammatory, anti-tumor, radiation protection, and scar removal both in vitro and in vivo. Full article

In adults, scars are formed after deep skin wound injuries like burns. However, the fetal microenvironment allows for scarless skin regeneration. One component that is abundantly present in the fetal extracellular matrix is hyaluronan (HA). To study whether biomaterials with HA improve wound healing, type I collagen scaffolds with and without HA were prepared and characterized. Their immune effect was tested using macrophages and their phenotypes were analyzed through cell surface markers and cytokine expression after 48 h. Since fibroblasts are the main cellular component in the dermis, adult, fetal and eschar-derived cells were cultured on scaffolds for 14 days and evaluated using histology, gene and protein expression analyses. Biochemical assays demonstrated that HA was successfully incorporated and evenly distributed throughout the scaffolds. Macrophages (M0) cultured on Col I+HA scaffolds exhibited a profile resembling the M2c-like phenotype (CD206^high, CD163^high and IL10^high). HA did not significantly affect gene expression in adult and fetal fibroblasts, but significantly reduced scarring-related genes, such as transforming growth factor beta 1 (TGFB1) and type X collagen alpha 1 chain (COL10A1), in myofibroblast-like eschar cells. These findings highlight the potential of incorporating HA into collagen-based skin substitutes to improve the wound healing response. Full article

Severe skin damage poses a significant clinical challenge, as limited availability of skin donors, postoperative skin defects, and scarring often impair skin function. Traditional two-dimensional (2D) nanofibers exhibit small pore sizes that hinder cellular infiltration, unable to simulate the three-dimensional (3D) structure of the skin. To address these issues, we developed 3D porous nanofiber scaffolds composed of polycaprolactone–polylactic acid–mussel adhesive protein (PLGA-PCL-MAP) using low-temperature electrospinning combined with nano-spray technology. Meanwhile, this 3D scaffold features high porosity, enhanced water absorption, and improved air permeability. The incorporation of mussel adhesive protein (MAP) further increased the scaffold’s adhesive properties and biocompatibility. In vitro experiments demonstrated that the 3D nanofiber scaffolds significantly promoted the adhesion, proliferation, and migration of epidermal keratinocytes (HaCaTs) and human fibroblasts (HFBs), while providing ample space for inward cellular growth. Successful co-culture of HaCaT and HFBs within the scaffold revealed key functional outcomes: HaCaTs expressed keratinocyte differentiation markers CK10 and CK14, while HFBs actively secreted extracellular matrix components critical for wound healing, including collagen I, collagen III, and fibronectin. This skin substitute with a composite structure of epidermis and dermis based on three-dimensional nanofiber scaffolds can be used as an ideal skin replacement and is expected to be applied in wound repair in the future. Full article

Objective: This study aims to evaluate the application effect of endoscopic thyroidectomy via the gasless unilateral axillary (GUA) approach in thyroid cancer and its impact on the postoperative stress response. Methods: Ninety-four thyroid cancer patients were enrolleod and assigned into the open group (underwent conventional-open-anterior-cervical-approach thyroidectomy) and the endoscopic group (underwent GUA endoscopic thyroidectomy) (n = 47). Perioperative indicators between the two groups were compared. Thyroid function parameters [total triiodothyronine (TT3), total thyroxine (TT4), free triiodothyronine (FT3), free thyroxine (FT4), and thyroid-stimulating hormone (TSH)] were measured preoperatively and on postoperative day 2. Inflammatory markers [interleukin-6 (IL-6) and tumor necrosis factor-α (TNF-α)] and stress-related hormones [norepinephrine (NE) and cortisol (Cor)] were evaluated preoperatively and on postoperative day 1. The aesthetic appearance of the incision was evaluated at 1 and 3 months postoperatively using the Vancouver Scar Scale (VSS). Postoperative complications were also compared between the two groups. Results: The endoscopic group exhibited less intraoperative blood loss, reduced postoperative drainage, a lower pain degree on 1 day postoperatively, a shorter hospitalization time, and a longer surgical time versus the open group (p < 0.05). The serum levels of TT3, TT4, FT3, and FT4 were lower, while the TSH levels were higher in both groups on postoperative day 2 compared to preoperative values. Additionally, the serum levels of IL-6, TNF-α, NE, and Cor increased on day 1 postoperatively, with the endoscopic group showing lower levels of these markers compared to the open group (p < 0.05). The VSS scores at 1 and 3 months after surgery were lower in the endoscopic group compared to the open group, indicating better cosmetic outcomes (p < 0.05). The incidence of postoperative complications was comparable between the endoscopic and open groups (p > 0.05). Conclusions: Endoscopic thyroidectomy by a GUA offers notable advantages over the conventional-open-anterior-cervical-approach thyroidectomy, including reduced intraoperative blood loss, less postoperative drainage, and a lower postoperative stress response. This approach also results in improved cosmetic outcomes, making it a promising alternative for thyroid cancer surgery. Full article

Background/Objectives: Cesarean scar pregnancy (CSP) refers to a pregnancy implanted on or within a scar from a previous cesarean birth. This study aims to evaluate the treatment strategies for CSP conducted at a single center and analyze the predictive markers of a single procedure without complications. Methods: A retrospective study was performed on CSP patients who received treatment at Korea University Ansan Hospital from November 2002 to December 2022. The clinical characteristics of CSP and the occurrence of complications based on treatment methods were investigated. Results: A total of 128 patients were included in the study. Among them, 84 patients (65.6%) underwent dilatation and curettage (D&C) only, 12 patients (9.4%) received a combination of methotrexate and D&C, 21 received D&C with Foley catheter ballooning (15.7%), and 11 patients (9%) underwent invasive procedures such as uterine artery embolization, hysterectomy, and open resection. As gestational age increased, the likelihood of opting for invasive procedures (aOR = 3.52, 95% CI 1.74–10.14, p = 0.003) also increased. A total of 84 patients (65.6%) were successfully treated with D&C without transfusion, and it was found that early gestational age and sonographic hypervascularity played a significant role. Conclusions: Early gestational age, younger maternal age, and sonographic hypervascularity were independently associated with favorable treatment without adjuvant therapy and transfusion. Therefore, for a safe and bloodless approach to treatment in cases of CSP, it is advisable to promptly diagnose this condition as early as possible. Full article

Family poultry production systems (FPPSs) in Chile and Argentina produce high-quality and nutritious food. However, little is known about chicken coccidiosis in these production systems. This work aimed to determine the Eimeria sp. positivity rate, circulating species, general farm management, and knowledge of the disease in FPPSs by gaining access to peri-urban markets in these two countries. The overall Eimeria sp. positivity rate, determined in 88 fecal samples, was 85.1%. Oocysts per gram in Argentinean meat-producing FPPSs were significantly higher than in Argentinean and Chilean egg-producing FPPSs. Multiplex-PCR based on seven Eimeria spp. Sequence-Characterized Amplified Region (SCAR) markers showed a great diversity of circulating species, with E. mitis (70.3%), E. acervulina (62.2%), and E. tenella (59.5%), followed by E. maxima (43.2%), E. praecox (32.4%), E. necatrix (18.9%), and E. brunetti (5.4%). Low awareness among family producers and low implementation of control measures were found. Importantly, pathogenic species were found even in asymptomatic chickens, which represents a potential chicken health threat. Furthermore, the administration of sulfonamides to broilers with clinical signs poses a risk of environmental contamination. This is the first comprehensive cross-sectional study showing that Eimeria sp. is a persistent parasite in peri-urban FPPSs in Argentina and Chile. Full article

Cutaneous lupus erythematosus (CLE) is a chronic autoimmune skin disorder with limited therapeutic options, particularly for refractory discoid lupus (DLE), which often results in scarring and atrophy. Recent studies have identified miR-31, miR-485-3p, and miR-885-5p as key regulators of inflammation, apoptosis, and fibrosis in CLE skin lesions. This research investigates a novel topical miRNA therapy using DDC642 elastic liposomes to target these pathways in CLE. DDC642 liposomes were complexed with miRNAs (anti-miR-31, anti-miR-485-3p, pre-miR-885-5p) and characterized through dynamic light scattering and Cryo-TEM. Cytotoxicity, cellular penetration, and therapeutic efficacy were evaluated in primary keratinocytes, PBMCs, and immune 3D-skin organoids. miRNA lipoplexes were successfully synthesized with optimized particle size, surface charge, and encapsulation efficiency. These lipoplexes exhibited effective cellular penetration and low cytotoxicity. Anti-miR-31 lipoplexes reduced miR-31 and NF-κB levels while increasing STK40 and PPP6C expression. Pre-miR-885-5p lipoplexes elevated miR-885-5p levels and downregulated PSMB5 and NF-κB in keratinocytes. While anti-miR-485-3p lipoplexes reduced T-cell activation markers. Anti-miR-31 and pre-miR-885-5p lipoplexes successfully modulated inflammatory pathways in 3D-skin CLE models. miRNA lipoplexes represent promising candidates for pioneering topical genetic therapies for CLE. Further studies, including animal models, are necessary to validate and optimize these findings. Full article

Fibrosis after full-thickness wound healing—especially after severe burn wounds—remains a clinically relevant problem. Biomaterials that mimic the lost dermal extracellular matrix have shown promise but cannot completely prevent scar formation. We present a novel approach where porous type I collagen scaffolds were covalently functionalized with ReGeneRating Agent (RGTA^®) OTR4120. RGTA^® is a glycanase-resistant heparan sulfate mimetic that promotes regeneration when applied topically to chronic wounds. OTR4120 is able to capture fibroblast growth factor 2 (FGF-2), a heparan/heparin-binding growth factor that inhibits the activity of fibrosis-driving myofibroblasts. Scaffolds with various concentrations and distributions of OTR4120 were produced. When loaded with FGF-2, collagen–OTR4120 scaffolds demonstrated sustained release of FGF-2 compared to collagen–heparin scaffolds. Their anti-fibrotic potential was investigated in vitro by seeding primary human dermal fibroblasts on the scaffolds followed by stimulation with transforming growth factor β1 (TGF-β1) to induce myofibroblast differentiation. Collagen–OTR4120(-FGF-2) scaffolds diminished the gene expression levels of several myofibroblast markers. In absence of FGF-2 the collagen–OTR4120 scaffolds displayed an inherent anti-fibrotic effect, as the expression of two fibrotic markers (TGF-β1 and type I collagen) was diminished. This work highlights the potential of collagen–OTR4120 scaffolds as biomaterials to improve skin wound healing. Full article