Down-regulation Of Gene Expression Research Articles

Pathomic and bioinformatics analysis of clinical-pathological and genomic factors for pancreatic cancer prognosis.

Pancreatic cancer exhibits a high degree of malignancy with a poor prognosis, lacking effective prognostic targets. Utilizing histopathological methodologies, this study endeavors to predict the expression of pathological features in pancreatic ductal adenocarcinoma (PAAD) and investigate their underlying molecular mechanisms. Pathological images, transcriptomic, and clinical data from TCGA-PAAD were collected for survival analysis. Image segmentation using unsupervised machine learning was employed to extract features, perform clustering, and establish models. The prognostic value of pathological features and associated clinical risk factors were evaluated; the correlation between pathological features and molecular mechanisms, gene mutations, and immune infiltration was analyzed. By clustering 45 effective pathological features, we divided PAAD patients into two groups: cluster 1 and cluster 2. Significant associations with poor prognosis were found for cluster 2 in both the training group (n = 113) and validation group (n = 75) (p = 0.006), with pathological stages II-IV identified as potential synergistic risk factors (HR = 2.421, 95% CI = 1.263-4.639, p = 0.008). Subsequently, through multi-omics correlation analysis, we further revealed a close association between cluster 2 and the oxidative phosphorylation mechanism. Within the cluster 2 group, 28 oxidative phosphorylation genes exhibited reduced expression, CDKN2A gene mutations were upregulated, and there was significant downregulation of Tregs infiltration and related immune gene expression. The pathomic model constructed using machine learning serves as a valuable prognostic target for PAAD. The histopathological features cluster 2 are closely associated with the downregulation of oxidative phosphorylation levels and Tregs immune infiltration.

Chlorogenic acid improves urogenital dysfunction induced by exposure to ambient particulate matter.

Oxidative stress is a well-known underlying mechanism for several diseases in response to environmental pollution. Although there is a lack of evidence on the relationship between air pollution and an established risk factor for urogenital dysfunction. The aim of this study was to investigate the mechanism of particulate matter (PM) on urogenital function and evaluate the potential efficacy of chlorogenic acid (CGA) in preventing urogenital damage in rats. Forty Wistar rats were divided into five groups (n = 8): control, particulate matter exposure (animals were exposed to fine dust in an inhalation chamber for 4 weeks, 3 days a week, for 3 h, PM10 concentration adjusted to 500-2000 µg/m3), and particulate matter plus 3 concentrations of chlorogenic acid (100, 200, and 400 mg/kg, gavage, 4 weeks, 3 days a week). At the end of the study, kidney biomarkers, oxidative stress markers, antioxidant enzymes, the oxidation resistance 1 (OXR1) and its downstream gene expression, sperm count, gonadotropin hormones, and the structure of the kidney, epididymis, and seminal vesicle were evaluated in response to PM exposure and CGA treatment in all groups. The data obtained from the current study showed that PM exposure led to kidney dysfunction and inhibition of oligospermia through oxidative stress, as evidenced by an increase in MDA and a decrease in TAC, SOD, CAT, and GSH concentration levels in blood samples. These results were consistent with the down-regulation of OXR1, Nrf2, and P21 gene expression. In contrast, CGA improved urogenital biomarkers and histopathology structures of the kidney, epididymis, and seminal vesicle by enhancing antioxidant defense system enzymes and modulating the OXR1 signaling pathway. Our findings suggest that environmental air pollution contributes to kidney dysfunction and urogenital damage. Modulation of oxidative stress through the OXR1, P21, and Nrf2 signaling pathways may be the underlying mechanism. Furthermore, chlorogenic acid supplementation could be recommended as a new protective or treatment strategy to safeguard urogenital function against exposure to particulate matter.

A microenvironment-modulating dressing with proliferative degradants for the healing of diabetic wounds.

Diabetic wounds are usually entangled in a disorganized and self-perpetuating microenvironment and accompanied by a prolonged delay in tissue repair. Sustained and coordinated microenvironment regulation and tissue regeneration are key to the healing process of diabetic wounds, yet they continue to pose a formidable challenge. Here we report a rational double-layered dressing design based on chitosan and a degradable conjugated polymer polydiacetylene, poly(deca-4,6-diynedioic acid) (PDDA), that can meet this intricate requirement. With an alternating ene-yne backbone, PDDA degrades when reacting with various types of reactive oxygen species (ROS), and more importantly, generates proliferative succinic acid as a major degradant. Inheriting from PDDA, the developed PDDA-chitosan double layer dressing (PCD) can eliminate ROS in the microenvironment of diabetic wounds, alleviate inflammation, and downregulate gene expression of innate immune receptors. PCD degradation also triggers simultaneous release of succinic acid in a sustainable manner, enabling long-term promotion on tissue regeneration. We have validated the biocompatibility and excellent performance of PCD in expediting the wound healing on both diabetic mouse and porcine models, which underscores the significant translational potential of this microenvironment-modulating, growth-promoting wound dressing in diabetic wounds care.

Utilizing the apical-out enteroids in vitro model to investigate intestinal glucose transport, barrier function, oxidative stress, and inflammatory responses in broiler chickens.

Conventional 2D intestinal epithelial cell lines have been widely used in investigating intestinal functions, yet with limitations in recapitulating the in vivo gut physiology of chickens. A recently established chicken enteroid model with apical-out nature and the presence of leukocyte components represents intestinal mucosal functions. The objectives of this study were to 1) evaluate basic gut nutrient transport and barrier functions in this model and 2) identify the model's effectiveness in studying inflammation and oxidative stress responses. Enteroids were generated from individual villus units isolated from the small intestine of Cobb500 broiler embryos. Enteroid viability, morphology, and epithelial cell markers were monitored; barrier function was evaluated based on the permeability to fluorescein isothiocyanate-dextran (FD4) with or without EDTA and lipopolysaccharide (LPS) challenges; nutrient transport was evaluated by fluorescence-labeled glucose (2NBD-G) with or without transporter blockade; the oxidative status was indicated by reactive oxygen species (ROS). Inflammatory and oxidative challenges were induced by LPS and menadione treatment, respectively. Selected marker gene expressions, including tight junction proteins (CLDN-1, CLDN-2, ZO-1, and OCCL), epithelial cell markers (Lgr-5, LYZ, and MUC-2), cytokines (IL-1β, IL-6, IL-8, IL-10, TNF-α, and INF-γ), and antioxidant enzymes (Nrf-2, catalase, and SOD), were determined by using RT-qPCR. Data were analyzed by one-way ANOVA among treatment groups. Enteroid cell activity was stable from day (d) 2 to d 6 and declined at d 7. Epithelial cell marker and cytokine expressions were stable from d 4 to d 6. FD4 permeability was increased after the EDTA treatment (P ≤ 0.05). Transporter-mediated 2NBD-G absorption was observed, which was reduced with glucose transporter blockade (P ≤ 0.05). Enteroids showed classic responses to LPS challenges, including upregulated gene expressions of IL-1β and IL-6, downregulated gene expressions of ZO-1 and OCCL, and increased FD4 permeability (P ≤ 0.05). Enteroids showed increased ROS generation (P ≤ 0.05) in response to oxidative stress. In conclusion, this apical-out enteroid model is a stable alternative in vitro model that exhibits intestinal barrier, nutrient transport, oxidation, and inflammation functions. With this enteroid model, we developed two challenge protocols for evaluating intestinal functions under oxidative stress and inflammation conditions.

Gene-Based Targeting for Treatment of Hypercholesterolemia and Prevention of Atherosclerotic Cardiovascular Disease

Familial hypercholesterolemia, characterized by high levels of LDL cholesterol is a monogenic metabolic disorder linked to atherosclerotic cardiovascular disease. Several genetic mutations in genes such as LDLR and PSK9 are thought to cause familial hypercholesterolemia. This ultimately causes dysfunction of the LDLR pathway, increasing circulating LDL levels. Current treatment revolves around decreasing LDL cholesterol levels in the blood. Statins are the current gold standard while sterol absorption inhibitors (ezetimibe) and PCSK9 inhibitors (evolocumab) are either used as primary treatment in individuals with statin intolerance, or supplementary treatment otherwise. However, CRISPR-Cas9 offers an upstream therapeutic intervention, inducing mutations in pathogenic genes (such as LDLR) to upregulate or downregulate gene expression and subsequently reduce LDL cholesterol levels. This technology thus has great potential as a two-fold treatment option, offering an effective solution for familial hypercholesterolemia, while simultaneously diminishing the risk of atherosclerotic cardiovascular disease. A limitation to current research exists in the sole analysis of monogenic factors in genetic screening, thus polygenic, multifactorial analyses are required to assess more holistic treatment plans. Furthermore, there exist current limitations to CRISPR-Cas9 including ethical limitations and social considerations revolving around the perpetuation of social inequities through technology necessitating future research on global health policies.

ELF1 serves as a potential biomarker for the disease activity and renal involvement in systemic lupus erythematosus

Systemic lupus erythematosus (SLE) is an autoimmune disease that affects multiple organs, yet its underlying mechanisms remain unclear, and precise biomarkers are lacking. In this study, we employed Mendelian randomization and HEIDI tools to comprehensively analyze large-scale Genome-Wide Association Study (GWAS) and expression Quantitative Trait Loci (eQTL) data, leading to the identification of seven novel potential functional genes associated with SLE, including BLK, ELF1, STIM1, B3GALT6, APOLD1, INPP5B, and FHL3. Subsequent investigations revealed a significant downregulation of ELF1 gene expression in CD4+ T cells of SLE patients compared to healthy controls. Moreover, within various SLE subgroups, such as those with decreased serum complement C3 levels, positive urinary protein, new-onset skin rashes, and SLE Disease Activity Index (SLEDAI) scores ≥ 5, ELF1 expression displayed a consistent decreasing trend. Notably, ROC curve analysis highlighted the diagnostic potential of ELF1 expression in SLE (AUC = 0.9493), as well as its value in assessing disease activity (AUC = 0.6852) and renal involvement (AUC = 0.7363). In conclusion, this study underscores the potential of ELF1 as a SLE biomarker for diagnosis and evaluation, offering insights into the underlying mechanisms of SLE and paving the way for future therapeutic research.

Replacement of fish meal with full fat Hermetia illucens modulates hepatic FXR signaling in juvenile rainbow trout (Oncorhynchus mykiss): exploring a potential role of ecdysteroids

The present study was conducted to investigate the effects of fish meal (FM) replacement with full fat Hermetia illucens (HI) on the molecular mechanisms regulating lipid and bile salt (BA) homeostasis in rainbow trout (Oncorhynchus mykiss) juveniles. We thus explore the presence of 20-hydroxyecdysone (20E) in an insect meal-based diet and evaluate its potential involvement in regulating the molecular mechanisms/basis of FXR:RXR axis signaling. Ecdysteroids are a category of steroid hormones which bind a nuclear–receptor complex composed of ecdysone receptor (EcR) and ultraspiracle protein (USP) and regulate insect molting and metamorphosis. In all vertebrates, including fish, EcR-USP homologs are the Farnesoid X receptors (FXR) and the Retinoid X receptors (RXR), which are known to regulate crucial physiological and metabolic aspects, including BA synthesis and cholesterol homeostasis. In silico prediction indicates that 20E binds the heterodimeric complex with a binding affinity constant Kd equals to 610±60 nM and affects positively the dimerization process. Results also demonstrated the coordinated increased expression of FXR and RXR, as well as their downstream target genes (i.e. short heterodimer partner 1 and 2) in rainbow trout fed diets containing HI meal. This latter finding was paralleled by a significant down-regulation of CYP7a1 and CYP8b1 gene expression together with a decrease in hepatic total cholesterol, triglyceride, and BA levels. Overall, our study suggested that FXR is a potential target for 20E content in insect meal and provided preliminary data on the potential role of ecdysteroids in regulating the metabolic status of teleost fish through modulation of FXR signaling in the enterohepatic system.

The CRISPR-dCas9 interference system suppresses inhA gene expression in Mycobacterium smegmatis

CRISPR-dead Cas9 interference (CRISPRi) has become a valuable tool for precise gene regulation. In this study, CRISPRi was designed to target the inhA gene of Mycobacterium smegmatis (Msm), a gene necessary for mycolic acid synthesis. Our findings revealed that sgRNA2 induced with 100 ng/ml aTc achieved over 90% downregulation of inhA gene expression and inhibited bacterial viability by approximately 1,000-fold. Furthermore, CRISPRi enhanced the susceptibility of M. smegmatis to isoniazid and rifampicin, which are both 50% and 90% lower than those of the wild-type strain or other strains, respectively. This study highlights the ability of CRISPRi to silence the inhA gene, which impacts bacterial viability and drug susceptibility. The findings provide valuable insights into the utility of CRISPRi as an alternative tool for gene regulation. CRISPRi might be further assessed for its synergistic effect with current anti-tuberculosis drugs and its possible implications for combating mycobacterial infections, especially drug-resistant tuberculosis.

Evolutionary divergent clusters of transcribed extinct truncated retroposons drive low mRNA expression and developmental regulation in the protozoan Leishmania

BackgroundThe Leishmania genome harbors formerly active short interspersed degenerated retroposons (SIDERs) representing the largest family of repetitive elements among trypanosomatids. Their substantial expansion in Leishmania is a strong predictor of important biological functions. In this study, we combined multilevel bioinformatic predictions with high-throughput genomic and transcriptomic analyses to gain novel insights into the diversified roles retroposons of the SIDER2 subfamily play in Leishmania genome evolution and expression.ResultsWe show that SIDER2 retroposons form various evolutionary divergent clusters, each harboring homologous SIDER2 sequences usually located nearby in the linear sequence of chromosomes. This intriguing genomic organization underscores the importance of SIDER2 proximity in shaping chromosome dynamics and co-regulation. Accordingly, we show that transcripts belonging to the same SIDER2 cluster can display similar levels of expression. SIDER2 retroposons are mostly transcribed as part of 3'UTRs and account for 13% of the Leishmania transcriptome. Genome-wide expression profiling studies underscore SIDER2 association generally with low mRNA expression. The remarkable link of SIDER2 retroposons with downregulation of gene expression supports their co-option as major regulators of mRNA abundance. SIDER2 sequences also add to the diversification of the Leishmania gene expression repertoire since ~ 35% of SIDER2-containing transcripts can be differentially regulated throughout the parasite development, with a few encoding key virulence factors. In addition, we provide evidence for a functional bias of SIDER2-containing transcripts with protein kinase and transmembrane transporter activities being most represented.ConclusionsAltogether, these findings provide important conceptual advances into evolutionary innovations of transcribed extinct retroposons acting as major RNA cis-regulators.

Gamma irradiation green synthesis of (polyacrylamide/chitosan/silver nanoparticles) hydrogel nanocomposites and their using as antifungal against Candida albicans and anti-cancer modulator

Silver nanoparticles-loaded hydrogel nanocomposites are exploited for medicinal and pharmaceutical applications. Hydrogel nanocomposites were prepared from acrylamide (Am), chitosan (CS) and AgNO3 utilizing gamma rays. Diverse variables were applied in preparation of silver nanoparticles-laoded hydrogel nanocomposites of (PAm/CS)-AgNPs such as influence of radiation dose and influnece of CS concentration. Diverse techniques were utilized to characterize hydrogel nanocomposites; Fourier transform infrared spectroscopy (FTIR), thermal gravimetric analysis (TGA), X-ray diffraction (XRD), Transmission electron microscopy (TEM), energy dispersive X-ray (EDX) and scanning electron microscopy (SEM). Results confirmed formation of silver nanoparticles-loaded hydrogel nanocomposites of (PAm/CS)-AgNPs. Antifungal activity of (PAm/CS)-AgNPs hydrogel nanocomposites on viability of C. albicans was esitmated. Results displayed the efficient microbial inhibition activity of treatment against C. albicans compared to control. Furthermore, (PAm/CS)-AgNPs hydrogel nanocomposite against cervical cancer HeLa cell line was investigated. Cytotoxicity of (PAm/CS)-AgNPs hydrogel nanocomposites on prior cancer cell line empolyed to prohibition of cell growth assesssed by MTT test. HeLa cancer cell is treated by (PAm/CS)-AgNPs for 48 h exposed a potential apoptotic activity by noticeable up-regulation of p53 gene expression. Moreover, anticancer activity was investigated by down-regulation of platelet-based growth variable receptor beta (PDGFR-β), Bcl2, Cathepsine, and MMP-2 gene expression. antioxidant activity was investigated and results showed antioxidant activity of (PAm/CS) hydrogel and (PAm/CS)-AgNPs hydrogel nanocomposite are 87.8% and 62.9%, respectively.

Statins have profound effects on the epicardial fat transcriptome

Abstract Background Epicardial adipose tissue (EAT) has direct contact with the myocardium, which enables a common microcirculation allowing bidirectional signaling between the tissues. It is possible that a change in the balance of protective and damaging mediators produced by EAT is a major driver for myocardial dysfunction. Notably, EAT volume and thickness associate with increased pro-inflammatory secretome and with coronary artery disease (CAD). Purpose This aim of this study was to analyze the total transcriptome in EAT and relate putative differences between patient groups to metabolic parameters as well as ongoing medications. Methods Between 2011 and 2016, 44 patients (mean age 68.0 years, 18.2% women) had a biopsy taken from the epicardial fat during coronoray by-pass surgery (CABG, n=36), or during elective aortic valvular replacement due to aortic stenosis (AVR, n=8). RNA from epicardial fat biopsis was extracted and sequenced. Differentially expressed genes were obtained by custom scripts that performed a negative binomial distribution test on effective counts. Multivariate regression analysis with orthogonal partial least-squares-discriminant analysis (OPLS-DA) was applied to extract and interpret the systematic variation in the resolved RNA sequencing profiles. To identify enrichment of biologically relevant functional gene sets, our data were entered in evidence-based network enrichment analysis (EviNet). Results The CABG surgery was performed approximately 7 days after an acute coronary event (75% acute MI), and 44% of them had previously known CAD. None of the AVR cases had significant CAD. At surgery, 97% of the CABG cases were prescribed statins, and 36% of them had statins alredy before the actual admission (long-term statin treatment). 63% of the AVR cases were on statins at admission. The OPLS analysis revealed only one medication with significant outcome, i.e., patients with long-term treatment with statins (n=18) versus patients without (n=26). Long-term statin treatment associated with down-regulated gene expression involved in beta-oxidation and triglyceride biosynthesis whereas gene expression involved in extracellular matrix organization were up-regulated. Gene expression involved in MHC class II presentation and interferon signalling were also reduced in patients with long-term treatment with statisn. A downregulation of genes in the interleukin- and complement cascade genes among CABG patient (versus AVR patients) without long-term statin treatment showed normal levels in statin pretreated patients. Conclusion Long-term statin treatment has a major influence on transcriptional regulation of genes involved in beta-oxidation, triglyceride biosynthesis, extracellular matrix composition, MHC class II presentation and interferon signalling in epicardial adipose tissue. These findings support important metabolic and immune effects of statins beyond the lipid-lowering function.

Construction of a tumor mutational burden-derived LncRNA prognostic computational framework associated with therapy sensitivity in skin cutaneous melanoma

BackgroundSkin cutaneous melanoma (SKCM) poses a significant public health challenge due to its aggressive nature and limited treatment options. To address this, the study introduces the Tumor Mutational Burden-Derived Immune lncRNA Prognostic Index (TILPI) as a potential prognostic tool for SKCM.MethodsTILPI was developed using a combination of gene set variation analysis, differential expression analysis, and COX regression analysis. Additionally, functional experiments were conducted to validate the findings, focusing on the effects of STARD4-AS1 knockdown on SKCM tumor cell behavior. These experiments encompassed assessments of tumor cell proliferation, gene and protein expression, migration, invasion, and in vivo tumor growth.ResultsThe results demonstrated that knockdown of STARD4-AS1 led to a significant reduction in tumor cell proliferation and impaired migration and invasion abilities. Moreover, it resulted in the downregulation of ADCY4, PRKACA, and SOX10 gene expression, as well as decreased protein expression of ADCY4, PRKACA, and SOX10. In vivo experiments further confirmed the efficacy of STARD4-AS1 knockdown in reducing tumor growth.ConclusionsThis study elucidates the mechanistic role of STARD4-AS1 and its downstream targets in SKCM progression, highlighting the importance of the ADCY4/PRKACA/SOX10 pathway. The integration of computational analysis with experimental validation enhances the understanding of TILPI and its clinical implications. Overall, the findings underscore the potential of novel computational frameworks like TILPI in predicting and managing SKCM, particularly through targeting the ADCY4/PRKACA/SOX10 pathway.

Bidirectional Impact of Varying Severity of Acute Kidney Injury on Calcium Oxalate Stone Formation

Introduction: Acute kidney injury (AKI) is a prevalent renal disorder. The occurrence of AKI may promote the formation of renal calcium oxalate stones by exerting continuous effects on renal tubular epithelial cells (TECs). We aimed to delineate the molecular interplay between AKI and nephrolithiasis. Methods: A mild (20 min) and severe (30 min) renal ischemia-reperfusion injury model was established in mice. Seven days after injury, calcium oxalate stones were induced using glyoxylate (Gly) to evaluate the impact of AKI on the formation of kidney stones. Transcriptome sequencing was performed on TECs to elucidate the relationship between AKI severity and kidney stones. Key transcription factors (TFs) regulating differential gene transcription levels were identified using motif analysis, and pioglitazone, ginkgetin, and fludarabine were used for targeted therapy to validate key TFs as potential targets for kidney stone treatment. Results: Severe AKI led to increased deposition of calcium oxalate crystals in renal, impaired kidney function, and upregulation of kidney stone-related gene expression. In contrast, mild AKI was associated with decreased crystal deposition, preserved kidney function, and downregulation of similar gene expression. Transcriptomic analysis revealed that genes associated with inflammation and cell adhesion pathways were significantly upregulated after severe AKI, while genes related to energy metabolism pathways were significantly upregulated after mild AKI. An integrative bioinformatic analysis uncovered a TF regulatory network within TECs, pinpointing that PKNOX1 was involved in the upregulation of inflammation-related genes after severe AKI, and inhibiting PKNOX1 function with pioglitazone could simultaneously reduce the increase of calcium oxalate crystals after severe AKI in kidney. On the other hand, motif analysis also revealed the protective role of STAT1 in the kidneys after mild AKI, enhancing the function of STAT1 with ginkgetin could reduce kidney stone formation, while the specific inhibitor of STAT1, fludarabine, could eliminate the therapeutic effects of mild AKI on kidney stones. Conclusion: Inadequate repair of TECs after severe AKI increases the risk of kidney stone formation, with the upregulation of inflammation-related genes regulated by PKNOX1 playing a role in this process. Inhibiting PKNOX1 function can reduce kidney stone formation. Conversely, after mild AKI, effective cell repair through upregulation of STAT1 expression can protect TEC function and reduce stone formation, and activating STAT1 function can also achieve the goal of treating kidney stones.

The DYRKP1 kinase regulates cell wall degradation in Chlamydomonas by inducing matrix metalloproteinase expression.

The cell wall of plants and algae is an important cell structure that protects cells from changes in the external physical and chemical environment. This extracellular matrix, composed of polysaccharides and glycoproteins, must be constantly remodeled throughout the life cycle. However, compared to matrix polysaccharides, little is known about the mechanisms regulating the formation and degradation of matrix glycoproteins. We report here that a plant kinase belonging to the DUAL-SPECIFICITY TYROSINE PHOSPHORYLATION-REGULATED KINASE (DYRK) family present in all eukaryotes regulates cell wall degradation after mitosis of Chlamydomonas reinhardtii by inducing the expression of matrix metalloproteinases (MMPs). Without the plant DYRK kinase (DYRKP1), daughter cells cannot disassemble parental cell walls and remain trapped inside for more than 10 days. On the other hand, the DYRKP1 complementation line shows normal degradation of the parental cell wall. Transcriptomic and proteomic analyses indicate a marked down-regulation of MMP gene expression and accumulation, respectively, in the dyrkp1 mutants. The mutants deficient in MMPs retain palmelloid structures for a longer time than the background strain, like dyrkp1 mutants. Our findings show that DYRKP1, by ensuring timely MMP expression, enables the successful execution of the cell cycle. Altogether, this study provides insight into the life cycle regulation in plants and algae.

Dynamic BMP gene expression regulation in chick RPE during recovery from short term optical defocus and form-deprivation.

This study investigated the differential gene expression of BMPs in chick retinal pigment epithelium (RPE) during recovery from short term exposure to optical defocus and form-deprivation (FD) treatments. 14-day old White-Leghorn chicks wore either monocular +10 or -10 D lenses, or diffusers for 2 or 48 h, after which eyes were allowed unobstructed vision for up to 96 h. Over this recovery period, refractive errors and choroidal thickness (ChT) were tracked using retinoscopy and high-frequency A-scan ultrasonography. Real-time PCR was used to examine the expression of BMP2, 4, and 7 genes in RPE samples collected 0, 15 min, 2, 24, 48, and 96 h after the termination of treatments. Expression levels in treated eyes and their contralateral control eyes were compared. After the termination of the lens and diffuser treatments, eyes gradually recovered from induced shifts in refractive error. With all three treatments, ChT changes reached statistical significance after 48 h of treatment, be it thinning with the -10 D lens and diffuser treatments (-0.06 ± 0.03mm, p < 0.05; -0.11 ± 0.04 mm, p < 0.05, resp.), or thickening with the +10 D lens (0.31 ± 0.04 mm, p < 0.001). BMP2 gene expression was rapidly upregulated in eyes wearing the +10 D lens, being statistical significance after 2 h, as well as 48 h of treatment. With the 2 h treatment, the latter gene expression pattern persisted for 15 min into the recovery period, before decreasing to the same level as that of contralateral control eyes, with a short-lived rebound, i.e., upregulation, 24 h into the recovery period. With the longer, 48 h treatment, BMP2 gene expression decreased more gradually, from 739 ± 121% at the end of the treatment period, to 72 ± 14% after 48 h of recovery. Two and 48 h of both -10 D and FD treatments resulted in BMP2 gene expression downregulation, with the time taken for gene expression levels to fully recover varying with the duration of initial treatments. In both cases, BMP2 gene expression downregulation persisted for 15 min into the recovery period, but reversed to upregulation by 2 h. Similar gene expression patterns were also observed for BMP4, although the changes were smaller. The observed changes in BMP gene expression in chick RPE imply dynamic, albeit complex regulation, with the duration of exposure and recovery being critical variables for all three types of visual manipulations. This study provides further evidence for a role of the RPE as an important signal relay linking the retina to the choroid and sclera in eye growth regulation.

Protective Effects of Lactobacillus Strains Against Oxidative Stress and Immune Suppression in Mice Receiving Aflatoxin-Contaminated Diet.

Mycotoxins like aflatoxins pose a significant threat to the health of both people and animals because of their deadly effects. This study aimed to investigate the potential of Lactobacillus strains in reducing the toxicity caused by aflatoxins in mice receiving a diet contaminated with aflatoxins. The mice were split up into various treatment groups, including a control group, an aflatoxin-treated group, and groups treated with the aflatoxin-contaminated diet along with Lactobacillus strains. Various parameters, including liver enzymes, blood parameters, malondialdehyde (MDA) levels, morphometric analysis of ileum, and gene expression, were analyzed to assess the effectiveness of the Lactobacillus strains in mitigating aflatoxins toxicity. Results showed that mice in the aflatoxin-treated group had increased MDA levels, indicating oxidative stress. Alternatively, the Lactobacillus cocktail treatment group showed a decreasing trend in MDA levels, suggesting a reduction in lipid peroxidation. The morphometric analysis of ileum tissue demonstrated that the Lactobacillus-treated group exhibited improved structural integrity compared to the aflatoxin-treated group. Additionally, gene expression analysis revealed that the Lactobacillus treatment attenuated the downregulation of SOD gene expression and mitigated the upregulation of iNOS gene expression induced by aflatoxins. These findings suggest that Lactobacillus strains have the potential to reduce aflatoxin-induced toxicity by alleviating oxidative stress, preserving intestinal tissue integrity, and modulating gene expression associated with antioxidant defense and inflammation. This study provides evidence for the beneficial effects of Lactobacillus strains in reducing aflatoxin toxicity in mice. The findings obtained may contribute to the development of preventive or therapeutic strategies against mycotoxin-induced toxicity.

The use of CRISPR-Cas9 technology for targeted gene therapy in Parkinson's disease focusing on LRRK2 and SNCA genes

Parkinson's disease (PD) is a neurodegenerative disorder characterized by the progressive degeneration of dopaminergic neurons in the substantia nigra pars compacta, leading to the start of motor and non-motor symptoms. These include bradykinesia, resting tremor, rigidity, and postural instability, alongside cognitive impairments, mood disorders, and autonomic dysfunctions. The hallmark of PD is the accumulation of α-synuclein protein aggregates, forming Lewy bodies within the neurons. Given the numerous causes of Parkinson’s, involving genetic, epigenetic, and environmental factors, therapeutic treatments are hard to come by. Traditional pharmacological treatments, such as levodopa and dopamine agonists, primarily offer symptomatic relief and are associated with diminishing efficacy over time. Therefore, there is a need for new therapeutic approaches that target the disease at its roots. CRISPR (Clustered Regularly Interspaced Short Palindromic Repeats) technology, with the RNA-guided Cas9 endonuclease enables precise, targeted modifications at the genomic level. This technology offers potential for terminating the genetic factors causing parkinsons and aid in developing gene-based therapies. By attempting to regulate gene expression, CRISPR holds promise for addressing the fundamental causes of neurodegeneration in Parkinsons. One of the key genetic targets in PD is the leucine-rich repeat kinase 2 (LRRK2) gene, where specific gain-of- function mutations, such as G2019S, lead to hyperactivation of its kinase activity, contributing to neurotoxicity. CRISPR-Cas9 can be employed to introduce precise double-strand breaks at the mutation site, followed by homology-directed repair (HDR) to restore the wild-type sequence, thereby normalizing LRRK2 function and preventing neuronal death. Similarly, CRISPR technology can target the SNCA gene, which encodes α-synuclein. Pathogenic duplicates or triplicates of SNCA result in overproduction of α-synuclein and subsequent aggregation. Utilizing CRISPR to disrupt these extra copies can reduce α-synuclein levels, alleviating its toxic effects.CRISPR activation (CRISPRa) and interference (CRISPRi) systems enable fine-tuned upregulation or downregulation of gene expression, offering strategies to increase the expression of neuroprotective factors or suppress the expression of deleterious genes.

Laminin-α2 chain deficiency in skeletal muscle causes dysregulation of multiple cellular mechanisms.

LAMA2, coding for the laminin-α2 chain, is a crucial ECM component, particularly abundant in skeletal muscle. Mutations in LAMA2 trigger the often-lethal LAMA2-congenital muscular dystrophy (LAMA2-CMD). Various phenotypes have been linked to LAMA2-CMD; nevertheless, the precise mechanisms that malfunction during disease onset in utero remain unknown. We generated Lama2-deficient C2C12 cells and found that Lama2-deficient myoblasts display proliferation, differentiation, and fusion defects, DNA damage, oxidative stress, and mitochondrial dysfunction. Moreover, fetal myoblasts isolated from the dy W mouse model of LAMA2-CMD display impaired differentiation and fusion in vitro. We also showed that disease onset during fetal development is characterized by a significant down-regulation of gene expression in muscle fibers, causing pronounced effects on cytoskeletal organization, muscle differentiation, and altered DNA repair and oxidative stress responses. Together, our findings provide unique insights into the critical importance of the laminin-α2 chain for muscle differentiation and muscle cell homeostasis.

A novel gene silencing strategy based on tobacco rattle virus in Hibiscus mutabilis.

Hibiscus mutabilis L. is a popular regional characteristic plant in China, cultivated for its attractive flower colors, extended bloom time, and medicinal properties. To enhance molecular breeding and gene function studies, we conducted transcriptome analysis and identified valuable genes in previous research. Nonetheless, the current inefficient and labor-intensive transformation techniques have hindered their applications. Virus-induced gene silencing (VIGS) provides a precise and effective strategy for post-transcriptional down-regulation of endogenous gene expression. We investigated the performance of tobacco rattle virus (TRV) as a tool for targeting and silencing the gene encoding the protein involved in chloroplast development, cloroplastos alterados 1 (altered chloroplast; CLA1), of H. mutabilis through Agrobacterium tumefaciens-mediated infiltration. By effectively suppressing the CLA1 gene associated with chloroplast development in H. mutabilis via the TRV-VIGS system, we have illustrated the inaugural implementation of VIGS in this species. Quantitative RT-PCR proved that HmCLA1 expression in agro-infiltrated plants was lower than in the mock-infiltrated (mock) and the control (CK) plants. Phenotypic observations corroborated the albino phenotype in leaves following successful HmCLA1 silencing. Our study showcases TRV-VIGS as a potential gene silencing tool for H. mutabilis, facilitating functional genomics studies and molecular breeding efforts in this species.

Heterogeneous Transcriptional Landscapes in Human Sporadic Parathyroid Gland Tumors

The expression of several key molecules is altered in parathyroid tumors due to gene mutations, the loss of heterozygosity, and aberrant gene promoter methylation. A set of genes involved in parathyroid tumorigenesis has been investigated in sporadic parathyroid adenomas (PAds). Thirty-two fresh PAd tissue samples surgically removed from patients with primary hyperparathyroidism (PHPT) were collected and profiled for gene, microRNA, and lncRNA expression (n = 27). Based on a gene set including MEN1, CDC73, GCM2, CASR, VDR, CCND1, and CDKN1B, the transcriptomic profiles were analyzed using a cluster analysis. The expression levels of CDC73 and CDKN1B were the main drivers for clusterization. The samples were separated into two main clusters, C1 and C2, with the latter including two subgroups of five PAds (C2A) and nineteen PAds (C2B), both differing from C1 in terms of their lower expression of CDC73 and CDKN1B. The C2A PAd profile was also associated with the loss of TP73, an increased expression of HAR1B, HOXA-AS2, and HOXA-AS3 lncRNAs, and a trend towards more severe PHPT compared to C1 and C2B PAds. C2B PAds were characterized by a general downregulated gene expression. Moreover, CCND1 levels were also reduced as well as the expression of the lncRNAs NEAT1 and VLDLR-AS1. Of note, the deregulated lncRNAs are predicted to interact with the histones H3K4 and H3K27. Patients harboring C2B PAds had lower ionized and total serum calcium levels, lower PTH levels, and smaller tumor sizes than patients harboring C2A PAds. In conclusion, PAds display heterogeneous transcriptomic profiles which may contribute to the modulation of clinical and biochemical features. The general downregulated gene expression, characterizing a subgroup of PAds, suggests the tumor cells behave as quiescent resting cells, while the severity of PHPT may be associated with the loss of p73 and the lncRNA-mediated deregulation of histones.