Search Results (132)

Fragile X syndrome (FXS, OMIM#300624) is the most common inherited cause of X-linked intellectual disability and behavior difficulties. In 99% of cases, it is caused by the pathological expansion (>200 repeats, full mutation -FM) of the CGG trinucleotide located at the 5′ UTR of the FMR1 (Fragile X Messenger Ribonucleoprotein 1) gene, leading to the lack of production of the FMRP. Clinical manifestations are well known in boys but are sometimes overlooked in girls, who may remain underdiagnosed. Premutation (PM) populations (55–200 repeats) may present other medical issues, such as FXPOI or FXTAS. Mosaic conditions, such as a combination of PM and FM lines in the same patient, may lead to milder phenotypes. With the improvement of genetic testing, information regarding the exact number of CGG triplet repeats and methylation status could help explain milder phenotypes in patients who may produce some FMRP. Chromosome X preferential inactivation (XCI) in FXS women can also play a role in clinical severity. We present four non-related families who were followed up in our FXS clinic. Some of their members showed FM on Southern blot, but had milder symptoms than expected. To rule out size mosaicism, a RT-PCR was performed, giving a different and more consistent molecular diagnosis. When mosaicism was not present, methylation status was performed, excluding full methylation. For females, XCI showed preferential inactivation in one case. Revisiting old molecular diagnoses should be considered in clinical practice, especially for patients with a milder phenotype than expected from their molecular reports. This personalized follow up may change their former diagnosis, prognosis, and expectations. Full article

Objective: We aimed to assess the expression and localization of renin-angiotensin system (RAS) receptors in lung tissue and the plasma concentration of related peptides in IPF patients. Materials and Methods: This case–control study involved 19 patients from southern Brazil undergoing lung resection or transplantation. Plasma levels of Angiotensin I, II, A, 1-7, Alamandine were measured via liquid chromatography–tandem mass spectrometry. Lung tissue expression and localization of angiotensin type 1 (AT1), Mas, and Mas-related G-protein-coupled receptor D (MrgD) receptors were evaluated using Western blot and immunohistochemistry. Clinical data and the 6-min walk test were analyzed to correlate receptor expression with lung function and oxygen dependence. Results: IPF patients showed reduced forced vital capacity (FVC) at 49 ± 13% and forced expiratory volume (FEV1) at 51 ± 14%, with a 60% increase in oxygen dependence. Plasma peptide concentrations were similar between the groups, except for Angiotensin I, which was significantly higher in the control group. In IPF lungs, AT1 and Mas receptors were expressed 2.31 and 2.13 times more, respectively, while MrgD expression was lower. Mas receptors were mostly found in bronchiole areas, whereas MrgD was predominant in the lung parenchyma. Conclusions: This study indicates that the RAS operates independently within tissue, in addition to its systemic functions, highlighting distinct differences between tissue and plasma RAS activities. The distinct roles of MrgD and Mas receptors in lung structure and function could be pivotal for new therapies, potentially leading to more effective IPF treatments. Full article

Fragile X syndrome (FXS) is the most common form of X-linked intellectual disability (ID). This study aimed to share 30 years of experience in diagnosing FXS and determine its frequency in Thailand. We retrospectively reviewed 1480 unrelated patients (1390 males and 90 females) with ID, developmental delay, or autism spectrum disorder, or individuals referred for FXS DNA testing at Songklanagarind Hospital, Thailand, over a 30-year period. The samples were analyzed using cytogenetic methods, PCR-based techniques, and/or Southern blot analysis. Full mutations (>200 CGG repeats) were identified in 100 males (7.2%) and three females (3.3%). An intermediate allele was detected in one male, while no premutation was found in the index cases. Two males were suspected to have FMR1 gene deletions. Twelve families underwent prenatal testing during this study. Most families undergoing prenatal FXS diagnosis involved mothers who were premutation carriers and had given birth to children affected by FXS. This study represents the largest series of molecular genetic FXS testing cases reported in Thailand. The frequency of FXS identified in different cohorts of Thai patients across various periods was approximately 7%. This study enhances public awareness of at-risk populations and highlights the importance of prenatal testing and genetic counseling for vulnerable families. Full article

Genetic transformation is an essential tool for investigating gene function and editing genomes. Kiwifruit, recognized as a significant global fresh fruit crop, holds considerable economic and nutritional importance. However, current genetic transformation techniques for kiwifruit are impeded by low efficiency, lengthy culture durations (a minimum of six months), and substantial labor requirements. In this research, we established an efficient system for shoot regeneration and the stable genetic transformation of the ‘Hayward’ cultivar, utilizing leaf explants in conjunction with two strains of Agrobacterium that harbor the expression vector pBI121-35S::GFP, which contains the green fluorescent protein (GFP) gene as a visible marker within the T-DNA region. Our results show that 93.3% of leaf explants responded positively to the regeneration medium, producing multiple independent adventitious shoots around the explants within a six-week period. Furthermore, over 71% of kanamycin-resistant plantlets exhibited robust GFP expression, and the entire transformation process was completed within four months of culture. Southern blot analysis confirmed the stable integration of GFP into the genome, while RT-PCR and fluorescence microscopy validated the sustained expression of GFP in mature plants. This efficient protocol for regeneration and transformation provides a solid foundation for micropropagation and the enhancement of desirable traits in kiwifruit through overexpression and gene silencing techniques. Full article

Nitrogen use efficiency remains the primary bottleneck for sustainable maize production. This study elucidates the functional mechanisms of the amino acid transporter ZmAAP1 in nitrogen absorption and stress resilience. Through systematic evolutionary analysis of 55 maize inbred lines, we discovered that the ZmAAP1 gene family exhibits distinct chromosomal localization (Chr7 and Chr9) and functional domain diversification (e.g., group 10-specific motifs 11/12), indicating species-specific adaptive evolution. Integrative analysis of promoter cis-elements and multi-omics data confirmed the root-preferential expression of ZmAAP1 under drought stress, mediated via the ABA-DRE signaling pathway. To validate its biological role, we generated transgenic maize lines expressing Arabidopsis thaliana AtAAP1 via Agrobacterium-mediated transformation. Three generations of genetic stability screening confirmed the stable genomic integration and root-specific accumulation of the AtAAP1 protein (Southern blot/Western blot). Field trials demonstrated that low-N conditions enhanced the following transgenic traits: the chlorophyll content increased by 13.5%, and the aboveground biomass improved by 7.2%. Under high-N regimes, the gene-pyramided hybrid ZD958 (AAP1 + AAP1) achieved a 12.3% yield advantage over conventional varieties. Our findings reveal ZmAAP1’s dual role in root development and long-distance nitrogen transport, establishing it as a pivotal target for molecular breeding. This study provides actionable genetic resources for enhancing NUE in maize production systems. Full article

Hemerocallis middendorffii is widely used in the landscaping of Northern China for its exceptional ornamental and ecological attributes. It is also the focus of a substantial body of germplasm development and stress tolerance research. However, the absence of an efficient regeneration and genetic transformation system has been a critical barrier to conducting gene function studies on this species. In this research, the aerial parts of seed-derived H. middendorffii plantlets were used as explants, and the callus induction, proliferation, subculture, differentiation, and rooting conditions in the in vitro regeneration process were optimized. A callus induction rate of 95.6% was achieved, with a regeneration rate of 84.4%. Based on this procedure, a simple and effective Agrobacterium-mediated genetic transformation system was preliminarily developed using a hygromycin-based selection system. The system comprised an Agrobacterium tumefaciens culture solution optical density at 600 nm (OD₆₀₀) of 0.6, an acetosyringone concentration of 100 μmol·L⁻¹ in both the A. tumefaciens infection solution and the co-cultivation medium, a sterilization culture with Timentin at 300 mg·L⁻¹, and a selection culture with hygromycin at 9 mg·L⁻¹. Transgenic H. middendorffii T0 rooted plants were produced within a 5-month period, with a transformation rate of 11.9% and positive rate of 32.8%. The regeneration and genetic transformation system established in this study should help advance functional gene research and genetic improvement in H. middendorffii. However, the genetic transformation was only validated in the T0 plants. To confirm stable integration and long-term transgene stability, future research on the phenotypic and molecular characterization of T1 progeny, including segregation analysis and Southern blot verification, will be conducted. Full article

Short tandem repeat (STR) sequences are highly variable DNA segments that significantly contribute to human neurodegenerative disorders, highlighting their crucial role in neuropsychiatric conditions. This article examines the pathogenicity of abnormal STRs and classifies tandem repeat expansion disorders(TREDs), emphasizing their genetic characteristics, mechanisms of action, detection methods, and associated animal models. STR expansions exhibit complex genetic patterns that affect the age of onset and symptom severity. These expansions disrupt gene function through mechanisms such as gene silencing, toxic gain-of-function mutations leading to RNA and protein toxicity, and the generation of toxic peptides via repeat-associated non-AUG (RAN) translation. Advances in sequencing technologies—from traditional PCR and Southern blotting to next-generation and long-read sequencing—have enhanced the accuracy of STR variation detection. Research utilizing these technologies has linked STR expansions to a range of neuropsychiatric disorders, including autism spectrum disorders and schizophrenia, highlighting their contribution to disease risk and phenotypic expression through effects on genes involved in neurodevelopment, synaptic function, and neuronal signaling. Therefore, further investigation is essential to elucidate the intricate interplay between STRs and neuropsychiatric diseases, paving the way for improved diagnostic and therapeutic strategies. Full article

Spain exhibits significant climatic variability across its regions, ranging from the humid oceanic climate in the north to the Mediterranean and stepped climates in the southern and central areas. These climatic differences influence environmental allergen exposure, which may, in turn, affect allergic sensitization patterns in the pediatric population. Variations in temperature, humidity, and airborne allergen distribution, such as pollen, dust mites, animal epithelia, and mold, contribute to regional disparities in allergic responses among children. Understanding how climatic conditions shape allergen recognition patterns across different geographical areas is essential for improving allergy prevention and management strategies. This study aims to shed light on this issue by identifying sensitization patterns in a pediatric population of 236 patients (with 2 age groups analyzed: 6–10 and 11–15 years old) from different climatic regions of Spain. Skin tests with standard aeroallergens were performed, and specific IgE (sIgE) analysis by Allergy Explorer of Macroarray Diagnostic test (ALEX²) and Western blot. The results revealed different sensitization trends across patients from the eight cities analyzed (Barcelona, Granada, Lugo, Sagunto, Santiago de Compostela, Valencia, and Zaragoza). Full article

We investigated the molecular and clinical profile of five boys carrying the fragile X messenger ribonucleoprotein 1 (FMR1) mutation and who suffered from the effects of prenatal alcohol exposure. Fragile X syndrome (FXS) testing was performed using PCR and Southern Blot analysis, and fragile X messenger ribonucleoprotein protein (FMRP) expression levels were measured by Western blot analysis. Clinical evaluation included cognitive functions, adaptive skills, autism phenotype, and severity of behavior measures. Fetal Alcohol Spectrum Disorder (FASD) was also assessed. Five adopted male siblings were investigated, four of which (cases 1, 2, 3, and 4) were diagnosed with FXS, FASD, and ASD, and one, the fraternal triplet (case 5), was diagnosed with FASD and ASD and no FXS. The molecular profile of case 1 and 2 showed the presence of a hypermethylated full mutation (FM) and the resulting absence of FMRP. Cases 3 and 4 (identical twins) were FM-size mosaics (for the presence of an FM and a deleted allele), resulting in 16% and 50% FMRP expression levels, respectively. FMRP expression level was normal in case 5 (fraternal twin). Severe behavioral problems were observed in all cases, including aggression, tantrum, self-harming, anxiety, and defiant behavior, due to different mutations of the FMR1 gene, in addition to biological exposure, home environmental factors, and potentially to additional background gene effects. Full article

ZDRF-8 is a transgenic maize event created via Agrobacterium-mediated transformation for insect resistance and glyphosate tolerance by expressing Cry1Ab, Cry2Ab, and G10evo-epsps. A Southern blot analysis suggested that it is a single-copy T-DNA insertion event. The flanking genomic sequences of the T-DNA insertion suggested that its T-DNA was inserted at the terminal region of the long arm of chromosome 7 without interrupting any known or predicted genes. Event-specific PCRs based on the flanking sequence were able to detect this event specifically. Laboratory bioassays and field trials of multiple generations demonstrated that ZDRF-8 is highly active against major corn pests in China, including Asian corn borers (ACB, Ostrinia furnacalis), cotton bollworms (CBW, Helicoverpa armigera), and oriental armyworm (OAW, Mythimna separata), and meanwhile confers glyphosate tolerance up to two times the recommended dose. The expression of the transgenes and the efficacy of insect resistance and glyphosate tolerance were stable over more than 10 generations. ZDRF-8 has been granted with a safety certificate in China, and its commercial release is expected in the coming years. Full article

Chios mastiha is the natural aromatic resin of Pistacia lentiscus L. var. Chia, Anacardiaceae, which is exclusively cultivated in the southern part of the Greek island of Chios. Chios mastiha (P. lenticonus/Chios mastiha) is well-known for its distinctive taste and aroma and has been known since ancient times due to its healing properties in gastrointestinal and inflammatory disorders and because of its anti-bacterial and anti-fungal activities. In this study, the chemical composition, applying LC-QTOF-MS/MS analysis, and the antioxidant activities of three different polarity P. lenticonus/Chios mastiha fractions, apolar, medium polar, and polar, were characterized in human neuroblastoma SH-SY5Y cells. Chemical analysis of the fractions unveiled new components of P. lenticonus/Chios mastiha, mainly fatty acids compounds, known for their antioxidant activity and regulatory effects on lipid metabolism. By applying the MTT assay and confocal microscopy analysis, we showed that P. lenticonus/Chios mastiha fractions, especially the apolar and medium polar fractions, enriched in triterpenes and fatty acids, caused suppression of the H₂O₂-induced reduction in cell viability, ROS production, and depolarization of the mitochondrial membrane potential, in SH-SY5Y cells. Moreover, Western blot analysis revealed that apolar fraction, enriched in fatty acids, induced expression of the PPARα, which is well-known for its antioxidant activities and its crucial role in lipid metabolism. Induction of PPARα, a GR target gene, was also accompanied by an increase in GR protein levels. Enhanced antioxidant activities of the apolar fraction may be correlated with its chemical composition, enriched in fatty acids and triterpenoids. Thus, our results indicate the neuroprotective actions of P. lenticonus/Chios mastiha fractions, highlighting their potential application as neuroprotective agents in neurodegenerative diseases. Full article

Peanut is a key cash crop worldwide, yet the limited availability of functional genes and markers for breeding hinders further progress, largely due to the lack of an efficient and user-friendly transformation system. This study aimed to comprehensively evaluate the effectiveness of nodal agroinjection, a novel transformation technique we developed for peanut, by introducing the soybean cold-tolerance gene SCTF-1. Putative transgenic seeds and seedlings were screened using genomic DNA PCR, while transgene expression was analyzed via qRT-PCR and phenotypic assessments. Southern blotting confirmed the stable integration of SCTF-1. The transgenic seedlings displayed enhanced chilling tolerance, characterized by increased proline accumulation, reduced malondialdehyde (MDA), and elevated peroxidase (POD) activity. These findings demonstrate that nodal agroinjection is an efficient and reliable approach for generating transgenic peanut and analyzing gene function. This method offers a promising alternative to conventional tissue culture-based transformation strategies. Full article

Torenia fournieri L. is a popular ornamental plant in the genus Torenia, widely used in commercial landscaping, especially during the summer. Additionally, Torenia has served as a model ornamental plant in many studies exploring ornamental characteristics and pest control through genetic engineering. To date, no research has been reported on developing insect-resistant Torenia expressing genes from Bacillus thuringiensis (Bt). In this study, a recombinant vector carrying the Cry1Ab gene from Bt, pBI121-Cry1Ab, was constructed and transferred into T. fournieri via Agrobacterium tumefaciens-mediated transformation. A total of 13 shoots survived on the kanamycin selection medium, among which four putative transgenic lines, designated L1, L2, L7, and L11, were molecularly confirmed by PCR and Southern blot analysis, indicating successful integration of the Cry1Ab gene into the genomes of these lines. Quantitative real-time PCR and ELISA results further verified the successful expression of the Cry1Ab gene in the leaves of all four transgenic lines. Insect bioassay results demonstrated that all four transgenic lines showed strong resistance to the insect pest, Mythimna separata, with mortality rates ranging from 59.9% to 100.0%, in contrast to a larval mortality rate of 16.2% in the wild-type Torenia. Additionally, these transgenic lines significantly decreased in larval survival rates compared to those fed on wild-type plants. Furthermore, these transgenic lines activated superoxide dismutase (SOD) activity at 12 and 24 h, and catalase (CAT) activity at 72 h, while suppressing SOD activity at 72 h, and peroxidase (POD) activity over time. Our findings indicate that these transgenic lines exhibit high resistance to the insect pest and provide new insights into controlling insect pests in ornamental plants through genetic approaches. Full article

Hepatitis B virus (HBV) can cause chronic infections, significantly increasing the risk of death from cirrhosis and hepatocellular carcinoma (HCC). A key player in chronic HBV infection is covalently closed circular DNA (cccDNA), a stable episomal form of viral DNA that acts as a persistent reservoir in infected hepatocytes and drives continuous viral replication. Despite the development of several animal models, few adequately replicate cccDNA formation and maintenance, limiting our understanding of its dynamics and the evaluation of potential therapeutic interventions targeting cccDNA. In this study, we aimed to develop a mouse model to investigate cccDNA formation and maintenance. We infected C57BL/6 mice with recombinant adeno-associated virus (rAAV) carrying a 1.3-overlength HBV genome (genotype C) and collected liver tissue at various time points to assess cccDNA levels and viral replication. Our results demonstrated the successful establishment of a chronic hepatitis B mouse model using rAAV-HBV1.3, which supported persistent HBV infection with sustained cccDNA expression in hepatocytes. Serum levels of HBsAg and HBeAg were elevated for up to 12 weeks, while alanine transaminase (ALT) levels remained within the normal range, indicating limited liver damage during this period. We confirmed HBV DNA expression in hepatocytes, and importantly, cccDNA was detected using qPCR after Plasmid-Safe ATP-Dependent DNase treatment, which selectively removes non-cccDNA forms. Additionally, Southern blot analysis confirmed the presence of cccDNA isolated using the Hirt extraction method. This established model provides a valuable platform for studying the long-term maintenance of cccDNA in chronic HBV infection and offers an important tool for testing novel therapeutic strategies aimed at targeting cccDNA. Full article

The herb Curculigo capitulata (Lour.) Ktze is widely distributed in southern and southwestern China. The Curculigo genus and its primary chemical constituents exhibit remarkable antidepressant activities. To investigate the chemical constituents and potential health benefits of C. capitulata, a phytochemical study was conducted. In this study, seven new compounds (capitugenin A–G), including three new norlignans (1–3), a new chalcone dimer (4), a new hemiacetal (5), two novel pyrrolidine-based compounds (6 and 7), including one identified as a natural product (7), and nineteen known compounds (8–26), were isolated from C. capitulata. The chemical structures and absolute configurations of Compounds 1–7 were elucidated via comprehensive spectroscopic data analyses. The neuroprotective effects of Compounds 1–26 against glutamate-induced cell death were tested in the human neuroblastoma cell line SH-SY5Y. Compounds 1, 3, 6, 8, 11, and 17 showed significant neuroprotective effects, with protection rates ranging from 29.4 to 52.8% at concentrations ranging from 5 to 40 μM. Western blot analysis indicated that Compound 3 exerted a protective effect by regulating the expression of Nrf2/HO-1. Full article