Search Results (259)

Background/Objectives: Human adenovirus 55 (HAdV55) is a notable pathogen causing community-acquired pneumonia; outbreaks occur frequently in military camps, hospitals, and schools, thereby posing a threat to public health security. This study aimed to develop a method for detecting HAdV55 nucleic acid by targeting the conserved region of the Hexon gene. The sequence was amplified using enzymatic recombination isothermal amplification (ERA) technology, in conjunction with CRISPR-Cas12a technology, to enhance the amplification signal. Methods: Optimized primer and crRNA sequences were selected through ERA isothermal amplification testing. The ERA-CRISPR/Cas12a detection method was completed within 30 min at a constant temperature of 42 °C. Results: Sensitivity was assessed by detecting standard plasmids and live strains at various dilution concentrations. The detection limits were determined to be 9 copies/reaction for standard plasmids and 2.5 copies/reaction for cultured HAdV55 strains. Specificity tests were conducted on positive samples for five common respiratory pathogens and five other adenovirus subtypes, all of which showed no cross-reactivity. Conclusions: A rapid ERA-CRISPR/Cas12a nucleic acid detection method for HAdV55 has been successfully developed, demonstrating high sensitivity and specificity without the need for expensive or complex instruments. This method holds promise for on-site pathogen screening and detection. Full article

Background: β-casomorphin-10 (CM-10), a peptide fragment derived from milk casein with the sequence YPFPGPIPNS, has demonstrated notable anxiolytic activity in BALB/c mice. Yet, its cellular responses and mechanistic pathways remain largely uncharacterized. Methods: We performed RNA-seq analysis to profile gene expression changes in δ-opioid receptor-expressing HEK293 cells (DOR-HEK), comparing CM-10-treated and untreated conditions. Results: CM-10 exposure led to differential expression of 1714 genes in DOR-HEK cells, with 34 upregulated (>1.4-fold) (1.9%) and 1680 downregulated (<0.71-fold) (98.1%), based on a predicted p-value threshold of <0.05. Notably, we identified 10 clusters that were associated with reduced cyclic AMP (cAMP) in DOR-HEK cells following CM-10 treatment. These clusters particularly involved genes related to regulatory subunits of cAMP-dependent protein kinases, such as PRKAR2A, cAMP-responsive element-binding pathway, circadian rhythms, such as CLOCK, ARNT1, CRY2, PER1, and PER2, and anxiety and depression, such as NOTCH1, NOTCH2 and ANK2. A network with these selected genes was confirmed by STRING analysis. Conclusions: These findings indicate that CM-10 may activate DOR-mediated signaling by suppressing cAMP levels, implicating a distinct molecular cascade in HEK293 cells. Full article

This study investigated the effect of lignite addition on maturity acceleration and bacterial community assembly mechanisms through high-throughput sequencing and iCAMP null model analysis in Co-composting of goat manure and corn straw. Three treatments were compared: 0% (CK), 5% (T1), and 10% (T2) lignite amendments (based on total solids). Results demonstrated that the 10% lignite addition significantly enhanced composting efficiency: the peak temperature increased by 0.83 °C, nitrogen loss and biotoxicity were reduced, the bacterial community structure was improved with elevated diversity and enriched keystone taxa, and the GI value was enhanced by 68.48 ± 2.99%. Notably, the relative abundances of key species, including Acinetobacter_indicus, Thermobacillus_composti, Pseudomonas_flexibilis, and Chelatococcus_thermostellatus, showed a significant positive correlation with the lignite addition gradient. The analysis of the symbiotic network centered on core microorganisms revealed that T2 showed the highest network complexity (254 links and 175 nodes), which was 46.8% and 48.3% higher than CK, respectively. Cooperative interactions dominated T2 networks, evidenced by a 13% increase in positive links for Acinetobacter (reaching 51.16%) and strengthened associations between humification-related genera (Chelatococcus and Thermobacillus links increased 1.4- and 3.2-fold, respectively). Using iCAMP null modeling, we further quantified assembly mechanisms: lignite addition increased stochastic processes (dispersal limitation increased from 47.57% in CK to 56.52% in T2) while reducing deterministic selection (homogeneous selection decreased from 25.57% to 18.47%). Acinetobacter, Chelatococcus, Ureibacillus, and Thermobacillus exhibited significant responsiveness to these ecological shifts. Collectively, 10% lignite amendment improved co-composting of goat manure and corn straw by synchronously accelerating maturity and restructuring bacterial assembly, providing a practical strategy for manure management via microbial regulation. Full article

Background: The health and viability of captive elephants, which are central to off-site conservation efforts and health management in Thailand, is threatened by emerging infectious diseases. This is partly due to genetic differences in immune-related genes, especially in the major histocompatibility complex (MHC) and, among these, loci such as DQA play a crucial role in immune surveillance. Data pertaining to MHC polymorphisms in elephants are scarce, and thus this study investigated such polymorphisms and selection signatures in a partial fragment of exon 2 of the MHC Class II DQA gene. Methods: The approach we used targeted next-generation sequencing and diversity analyses of individuals from three captive elephant camps in Northern Thailand. Results: Eight alleles containing 11 SNPs were identified in the exon 2 fragment, encompassing both silent and missense mutations, some of which may influence immune function. Notably, the allele Elma-DQA*TH3, which is identical to Loaf-DQA*01 and Elma-DQA*01, previously reported as the most common alleles in Loxodonta and Elephas, was found at low frequencies. This shift may reflect local selective pressures that shape MHC allele distributions. Evidence of mixed selection (both positive and balancing) was detected in the partial fragment of DQA exon 2, suggesting a dynamic interplay between evolutionary forces. Positive selection likely reflects an adaptation to emerging or locally prevalent pathogens, whereas balancing selection maintains allelic diversity over time to enable a broad immunological response. Conclusions: Our findings reveal immunogenetic variations in captive Thai elephants, and provides insights into host–pathogen interactions that inform conservation and health strategies with the aim of improving disease resilience. Full article

Hydroethanolic Cucurbita pepo seed extracts are traditionally used for alleviating lower urinary tract symptoms (LUTS), yet their mechanisms remain unclear. Adenosine, a purine nucleoside involved in neuromodulation and smooth muscle relaxation, was recently identified in C. pepo seeds. Since A₁ adenosine receptors (A₁AR) suppress parasympathetic bladder overactivity by inhibiting acetylcholine (ACh) release, we investigated to which extent purines from pumpkin seed extracts contribute to A₁AR activation. Complementary antioxidant capacity was assessed using the 2,2-diphenyl-1-picrylhydrazyl (DPPH) assay. Three hydrophilic seed extracts containing different adenosine levels (0.60–1.18 mg/g dw) were evaluated for agonist activity using a cAMP inhibition assay. The most active extract showed an EC₅₀ of 40.22 µg/mL. Selective removal of adenosine shifted the dose–response curve rightward, while further elimination of an adenosine derivative increased the EC₅₀ to 212.10 µg/mL, confirming adenosine as the principal active compound. Guanosine and inosine did not exhibit A₁AR agonist or allosteric effects. All samples exhibited measurable but weak antioxidant activity (IC₅₀ = 1.02–4.19 mg/mL), consistent with their low total phenolic content. These findings underscore the importance of accounting for naturally occurring agonists in plant extracts to avoid overestimating receptor-mediated effects in vitro which are not translatable in vivo. Full article

Since the first paper published by Susan Cole in 1990 detailing multidrug resistance mediated by ABCC1/MRP1, research into the C-subfamily of ATP-binding cassette transporters has continued to uncover a wide range of functionally divergent proteins. However, several orphan transporters remain in the C-subfamily, and the physiological function and substrates of ABCC5, ABCC11, and ABCC12 remain elusive. This review explores the emerging understanding of human ABCC5. Unlike other ABC transporters with well-defined drug export functions, ABCC5’s physiological roles remain only partially understood. While it is known for its involvement in multidrug resistance in cancers, recent studies suggest broader implications in development, metabolism, neurobiology, and male fertility. ABCC5 exports various endogenous substrates, including cyclic nucleotides (cAMP and cGMP), glutamate conjugates like NAAG, and possibly haem. Knockout models in mice, zebrafish, and sea urchins reveal ABCC5’s role in gut formation, brain function, eye development, and iron metabolism. In mice, its deletion results in lower adipose tissue mass, enhanced insulin sensitivity, and neurobehavioral changes resembling schizophrenia, highlighting its role in glutamatergic signalling and circadian regulation. Functionally, ABCC5 appears to impact adipocyte differentiation and GLP-1 release, implicating it in type 2 diabetes susceptibility in humans. Structural studies using human ABCC5 revealed a novel autoinhibitory mechanism involving a peptide segment (C46–S64) that blocks substrate binding, offering new potential for selective inhibitor development. However, this review emphasises caution in targeting ABCC5 for cancer therapy due to its underappreciated physiological function(s), particularly in the brain and male reproductive system. Understanding ABCC5’s substrate specificity, regulatory mechanisms, and functional redundancy with its paralog ABCC12 remains critical for successful therapeutic strategies in humans. Full article

The corticotropin-releasing factor (CRF) and its type 1 receptor (CRF₁R) play a key role in the regulation of the hypothalamic–pituitary–adrenal (HPA) axis. Dysregulation of the HPA axis is associated with congenital adrenal hyperplasia (CAH) and depression. Non-peptide CRF₁R-selective antagonists displayed antidepressant effects on animal models and are used for the management of CAH. To develop novel non-peptide CRF₁R antagonists, we have previously designed and synthesized a series of substituted pyrimidines. Among these analogs, molecule 43 (M43) binds to CRF₁R with the highest affinity. Based on this finding, we selected M43 for further pharmacological characterization in the present study. The results suggest that M43 is a potent CRF₁R antagonist, blocking the ability of the CRF-related agonist, Tyr⁰-sauvagine, to stimulate (1) cAMP accumulation in HEK 293 cells expressing CRF₁R and (2) the proliferation rate of RAW 264.7 macrophages. Computational studies suggest that the antagonist properties of M43 are mostly attributed to its ability to interact with residues in the allosteric pocket of CRF₁R, comprised of the third, fifth, and sixth transmembrane domain residues, which block activation-associated structural rearrangements of the receptor. Our data will be used to design novel non-peptide CRF₁R antagonists for clinical use. Full article

Background/Objectives: Metabolic/bariatric surgery is currently the most successful treatment for patients with obesity; however, a fifth of patients undergoing surgery may not lose enough weight to be considered successful. Recent studies have shown that bariatric/metabolic surgery alters the epigenome and may explain postoperative improvements in metabolic health. The primary objective is to consolidate published differentially methylated CpG sites in pre- and post-metabolic/bariatric surgery female patients and associate them with the respective genes and pathways. Methods: This systematic review adhered to the PRISMA-P guidelines and was registered with the PROSPERO (CRD42023421852). Following an initial screening of 541 studies using COVIDENCE, six studies were selected, comprising three epigenome-wide association studies (EWAS) and three candidate gene methylation studies. The published studies collected DNA samples from female patients with obesity before and after surgery (3 months, 6 months, 9–31 months, and 2 years). KEGG pathway analysis was performed on genes where the extracted CpG sites were located. Results: The meta-analysis showed that 11,456 CpG sites are differentially methylated after a successful weight loss surgery, with 109 sites mapped to genes involved in key metabolic pathways, including FoxO, mTOR, insulin, cAMP, adipocytokine, Toll-like receptor, and PI3K-Akt. Conclusion: The highlighted differentially methylated CpG sites can be further used to predict the molecular signature associated with successful metabolic/bariatric surgery. Full article

Background: This study investigates the expression of microRNAs (miRNAs) in the semitendinosus muscle of cattle breeds with varying intramuscular fat (IMF) deposition to identify key miRNA regulators of beef marbling, utilizing Hereford (HER; higher IMF) and Holstein-Friesian (HF; moderate IMF) bulls, and Limousin (LIM; low IMF) bulls with lower IMF in the semitendinosus muscle. Methods: MicroRNA profiling used custom bovine microarrays and the Agilent software. The selected miRNAs, miR-34a, miR-149-5p, miR-208b, miR-499, miR-660, and miR-1343-5p, were chosen for validation using real-time PCR, confirming their differential expression. Target prediction utilized miRWalk, while functional and pathway analyses were conducted using the DAVID database to interpret biological relevance. Results: Microarray analysis identified 51 differentially expressed miRNAs. Among these, 24 exhibited consistent expression patterns in high-marbling breeds compared to the low-marbling LIM breed. Bioinformatic analysis of the 4941 predicted target genes of these 24 miRNAs revealed significant enrichment in pathways crucial for marbling, including the adipocytokine, AMPK, MAPK, and PI3K-Akt signaling pathways, as well as biological processes such as cell differentiation and lipid homeostasis. Notably, miR-34a and miR-149-5p emerged as significant regulators, with miR-34a targeting genes like SIRT1, HMGA2, PTPN11, VEGFA, FGF1, FGF2, and BRAF, and miR-149-5p influencing adipogenesis and lipid metabolism through its association with crucial KEGG pathways such as PI3K–Akt, MAPK, PPAR, TGF-β, cAMP, and Wnt signaling, all of which collectively influence adipocyte differentiation, lipid metabolism, cell cycle control, and angiogenesis. Conclusions: The findings underscore identified miRNAs’ possible coordinated regulatory role, particularly miR-34a and miR-149-5p, in the complex molecular mechanisms governing IMF deposition in cattle, providing potential targets for improving beef quality. Full article

Articular cartilage (AC) has a very limited capacity for self-healing once damaged. Chondrocytes maintain AC homeostasis and are key cells in AC tissue engineering (ACTE). However, chondrocytes lose their function due to oxidative stress. Umbilical cord mesenchymal stem cells (UCMSCs) are investigated as an alternative cell source for ACTE. MSCs are known to regulate tissue regeneration through host cell modulation, largely via extracellular vesicle (EV)-mediated cell-to-cell communication. The purpose of this study was to verify whether UCMSC-derived EVs (UCMSC-EVs) enhance chondrocyte function. The mean particle sizes of the UCMSC-EVs were 79.8 ± 19.05 nm. Transmission electron microscopy (TEM) revealed that UCMSC-EVs exhibited a spherical morphology. The presence of CD9, CD63, and CD81 confirmed the identity of UCMSC-EVs, with α-tubulin undetected. UCMSC-EVs maintained chondrocyte survival, and increased chondrocyte proliferation after intake by chondrocytes. UCMSC-EVs upregulated mRNA levels of SOX-9, collagen type II (Col-II), and Aggrecan, while decreasing collagen type I (Col-I) levels. UCMSC-EVs reduced the oxidative stress of chondrocytes by reducing mitochondrial superoxide production and increasing protein levels of SOD-2 and Sirt-3 in chondrocytes. The 50 most abundant known microRNAs (miRNAs) derived from UCMSC-EVs were selected for gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses. GO analysis revealed enrichment in pathways associated with small GTPase-mediated signal transduction, GTPase regulatory activity, and mitochondrial matrix. The KEGG analysis indicated that these miRNAs may regulate chondrocyte function through the PI3K-Akt, MAPK, and cAMP signaling pathways. In summary, this study shows that UCMSC-EVs enhance chondrocyte function and may be applied to ACTE. Full article

Background: Signaling pathways like those depending on cAMP/PKA, calcium/calmodulin/CaMK, MEK-1/MAPK or PI3K/Akt have been described to modulate suprachiasmatic nucleus (SCN) neuronal signaling via influencing transcription factors like CREB. Here, we analyzed the effect of cyclic nucleotide phosphodiesterase inhibitors and structurally similar substances commonly used as autophagy modulators on a cell line stably expressing a cyclic nucleotide element-driven luciferase reporter. Methods: We used an SCN cell line stably transfected with a CRE-luciferase reporter (SCNCRE) to evaluate signaling and vitality responses to various isoform-selective PDE inhibitors and autophagy modulators to evaluate the mechanism of action of the latter. Results: In this study the different impacts of common PDE inhibitors and autophagy modulators on CRE-luciferase activity applied alone and in combination with known CRE-luciferase activating agents showed that (1) PDE3, 4 and 5 are present in SCNCRE cells, with (2) PDE3 being the most active and (3) the autophagy inhibitor 3-Methyladenin (3-MA) displaying PDE inhibitor-like behavior. Conclusions: Experiments provide evidence that, in addition to the extracellular signaling pathways components shown before to be involved in CRE-luciferase activity regulation like cAMP analogs, adenylate cyclase activators and beta-adrenoceptor agonists, cyclic nucleotide metabolism as realized by phosphodiesterase activity, or molecule/agents influencing processes like autophagy or inflammation, modulate transcriptional CRE-dependent activity in these cells. Specifically, we provide evidence that the autophagy inhibitor 3-MA, given that PDEs are expressed, may also act as a PDE inhibitor and inducer of CRE-mediated transcriptional activity. Full article

Spurred by the enormous therapeutic success of glucagon-like peptide-1 receptor (GLP-1R) agonists (GLP1-RAs) against diabetes and obesity, glucagon family receptor pharmacology has garnered a tremendous amount of interest. Glucagon family receptors, e.g., the glucagon receptor itself (GCGR), the GLP-1R, and the glucose-dependent insulinotropic peptide receptor (GIPR), belong to the incretin receptor superfamily, i.e., receptors that increase blood glucose-dependent insulin secretion. All incretin receptors are class B1 G protein-coupled receptors (GPCRs), coupling to the G_s type of heterotrimeric G proteins which activates adenylyl cyclase (AC) to produce cyclic adenosine monophosphate (cAMP). Most GPCRs undergo desensitization, i.e., uncouple from G proteins and internalize, thanks to interactions with the βarrestins (arrestin-2 and -3). Since the βarrestins can also mediate their own G protein-independent signaling, any given GPCR can theoretically signal (predominantly) either via G proteins or βarrestins, i.e., be a G protein- or βarrestin-“biased” receptor, depending on the bound ligand. A plethora of experimental evidence suggests that the GLP-1R does not undergo desensitization in physiologically relevant tissues in vivo, but rather, it produces robust and prolonged cAMP signals. A particular property of constant cycling between the cell membrane and caveolae/lipid rafts of the GLP-1R may underlie its lack of desensitization. In contrast, GIPR signaling is extensively mediated by βarrestins and the GIPR undergoes significant desensitization, internalization, and downregulation, which may explain why both agonists and antagonists of the GIPR exert the same physiological effects. Here, we discuss this evidence and make a case for the GLP-1R being a phenotypically or functionally G_s-selective receptor. We also discuss the implications of this for the development of GLP-1R poly-ligands, which are increasingly pursued for the treatment of obesity and other diseases. Full article

Background: The aim of this scoping review is to analyze the health status of Roma and Sinti in Europe, highlighting the issues faced by children and women. In addition, we want to examine the access of these groups to health care services and to identify possible interventions to increase their use. Methods: Our research was conducted on Pubmed, Google Scholar, and the Trip Database. We selected articles written in English, Spanish, and Italian published since 2015. Results: Studies have shown that the health status of Roma and Sinti populations is generally worse than that of the rest of the population. Limited access to care is due to several specific factors, such as beliefs, traditions, and the lack of awareness of widespread direct and indirect discrimination against these groups by healthcare professionals. The studies reviewed have shown how mistrust can be broken down through multi-centered interventions linked to information, education, and communication through mediators able to interact with these populations, as well as through appropriate training of the health workers in charge. Conclusions: The health of the Roma and Sinti populations is commonly worse than that of the rest of the population. This is particularly true for the large proportion of people confined to suburban camps. However, the available evidence signals the low quality of life they experience and the need for interventions involving the communities and the establishment of ad hoc orientation or initial care contact points in the segregated areas. This could lead to an improvement in the integration of this population into the National Health Systems’ activities. Full article

Phosphodiesterase 4 (PDE4) serves as a crucial regulator of cyclic adenosine monophosphate (cAMP) signaling and has been identified as a significant therapeutic target for inflammatory and metabolic disorders impacting the gastrointestinal (GI) tract and liver. Although pan-PDE4 inhibitors hold therapeutic promise, their clinical use has been constrained by dose-dependent adverse effects. Recent progress in the development of isoform-specific PDE4 inhibitors, such as those selective for PDE4B/D, alongside targeted delivery systems like liver-targeting nanoparticles and probiotic-derived vesicles, is reshaping the therapeutic landscape. This review consolidates the latest insights into PDE4 biology, highlighting how the structural characterization of isoforms informs drug design. We conduct a critical evaluation of preclinical and clinical data across various diseases, including inflammatory bowel diseases (IBDs), alcoholic liver disease, nonalcoholic fatty liver disease (NAFLD), liver fibrosis, and digestive tract tumors, with an emphasis on mechanisms extending beyond cAMP modulation, such as microbiota remodeling and immune reprogramming. Additionally, we address challenges in clinical translation, including biomarker discovery and the heterogeneity of trial outcomes, and propose a roadmap for future research directions. Full article

CB₁ and CB₂ cannabinoid receptors are members of the GPCR superfamily that modulate the effects of endocannabinoids. CB₁ is the most abundant CB receptor in the central nervous system, while CB₂ is present both peripherally and in the brain. CB₂ plays a role in inflammation, as well as neurodegenerative and psychiatric disorders. To identify new ligands for CB₂, we screened a library of FDA-approved drugs for activity at the receptor using a thallium flux assay, resulting in the discovery of the immunosuppressant mycophenolate mofetil as a potent, selective activator of CB₂. Further characterization of the compound confirmed agonist activity in a variety of complementary assays, including PI hydrolysis, cAMP inhibition, and β-arrestin recruitment. Radioligand binding assays established a non-competitive interaction with the site occupied by [³H]CP55,940. CB₂ agonists GW-842,166X and MDA7 were also profiled, revealing that GW-842,166X exhibits a similar activity profile to mycophenolate mofetil, whereas MDA7 presents a distinct profile. These differences provide insight into the complex CB₂ pharmacology impacting preclinical and clinical studies, and ultimately, new treatment strategies for brain disorders. Full article