Patient-reported outcome measures (PROMs) are important tools for assessing the impact of skin diseases on health-related quality of life (HRQoL). However, comparative data on the measurement properties of commonly used instruments are limited. The objective of this study was to compare the measurement properties of the Dermatology Life Quality Index (DLQI), DLQI-Relevant (DLQI-R), and Skindex-29 in a cohort of patients with chronic skin disease. Participants completed the DLQI, Skindex-29, and Patient-Reported Outcomes Measurement Information System (PROMIS) measures at baseline and 3 months. Structural validity was assessed with confirmatory factor analysis; internal consistency with Cronbach's alpha; construct validity via known-groups comparisons and PROMIS correlations; informativity with Shannon's index; and responsiveness with standardized response means, Cohen's d, and change score correlations. Of 549 participants (mean age 37.8 years; 52.3% female), 406 (74.0%) completed follow-up. All instruments demonstrated high internal consistency (α ≥ 0.85) and no floor or ceiling effects. Skindex-29 domains demonstrated stronger structural validity and informativity compared to DLQI and DLQI-R. Construct validity was supported by expected associations with patient global assessment (PtGA) and PROMIS correlations. The DLQI, DLQI-R, and Skindex Symptoms and Emotions showed moderate responsiveness. The DLQI, DLRI-R and Skindex-29 demonstrated strong measurement properties in a heterogeneous cohort of individuals with chronic skin disease. Skindex-29 showed stronger structural validity and informativity, while the DLQI and DLQI-R demonstrated slightly greater responsiveness. Selection of PROM should depend on the intended clinical or research purpose. Chronic skin conditions, such as acne, eczema, psoriasis, and hidradenitis suppurativa, can affect many areas of daily life, including symptoms, emotional wellbeing, and social interactions. Physicians and researchers often use questionnaires, called patient-reported outcome measures, to better understand this impact from the patient’s perspective. Two of the most widely used in dermatology are the Dermatology Life Quality Index (DLQI), and the Skindex-29. A newer scoring approach to the DLQI, known as the DLQI-Relevant (DLQI-R), was developed to improve accuracy by adjusting how responses are scored when patients indicate that certain questions do not apply to them. In this study, we wanted to understand which of these questionnaires works best in people living with chronic skin conditions. We surveyed over 500 adults in the United States who reported having one or more skin condition. The participants completed the DLQI and the Skindex-29 at the start of the study, and again 3 months later. We found that the Skindex-29 provided a broader and more detailed picture of how skin conditions affect health-related quality of life. The DLQI and DLQI-R, on the other hand, were slightly better at detecting changes in health-related quality of life over time, although the differences between the questionnaires were small. These results show that no single instrument is best for all situations. The choice of questionnaire should depend on the purpose—whether the goal is to capture the full burden of disease, or to measure how much a patient improves with treatment.
Medication-related osteonecrosis of the jaw (MRONJ) is a complex condition associated with the use of antiresorptive drugs, such as bisphosphonates and denosumab. The condition is characterized by the presence of exposed bone in the maxillofacial region that fails to heal. MRONJ remains highly intractable, as its pathogenic mechanisms are not yet fully understood. It is therefore essential to elucidate the molecular mechanisms underlying the disease. MiRNA expression analysis and proteomic studies were performed on a selected cohort of patients with MRONJ on jawbone tissue, using qRT-PCR and 2D electrophoresis followed by mass spectrometry. MiRNAs and proteomics data validation was carried out by Western blot analysis of differentially expressed proteins highlighted by a proteome study and predicted targets of differentially expressed miRNAs. Nineteen miRNAs were overexpressed and two downregulated in jawbone tissue from all MRONJ patients. Notably, five of these dysregulated miRNAs are involved in the regulation of angiogenesis and desmosome functions, suggesting a potential link to the molecular alterations observed at the protein level. Proteomic analysis revealed decreased concentrations of the pigment epithelium-derived factor, and of desmoglein-1, a desmosomal cadherin. Validation analysis confirmed the dysregulation of pathways involved in bone remodeling and necroptosis. The pathophysiology of MRONJ arises from a complex interplay of factors, including impaired bone remodeling, affected angiogenesis, and altered cell adhesion and differentiation mechanisms, ultimately leading to necroptosis. Through proteomic analysis and validation of miRNA expression, our study proposes specific molecular alteration in MRONJ-compromised bone tissue, involving desmosomal component imbalance and angiogenesis inhibition.
<p>Background. Patellofemoral instability is a multifactorial condition influenced by anatomical abnormalities as well as soft tissue dysfunction. Distalization of the tibial tuberosity is classically reserved for patients with patella alta, typically defined by a Caton-Deschamps (CD) index &gt;1.2. However, clinical experience suggests that certain patients with lower CD indices but distally deepening trochlear morphology may also benefit. This study aimed to evaluate the long-term outcomes of tibial tuberosity osteotomy with distalization and medialization (TTO-DM) in patients with recurrent lateral patellar instability, regardless of patellar height.<br />Materials and methods. This retrospective single-center study included 26 patients (17 females, 9 males) with recurrent patellar instability and Dejour type A, B, or C trochlear dysplasia. All demonstrated a distally deepened trochlear groove on MRI and underwent TTO-DM along with MPFL reconstruction. Patients were followed up for a minimum of 4 years (mean, 8.6 years). Outcome measures included the Kujala and Lysholm scores, radiographic assessment of the CD index and Kellgren-Lawrence grade, complications, range of motion, and pain during squatting.<br />Results. Kujala scores improved from 60.1 18.7 to 93.8 7.1 (p &lt; 0.0001), and Lysholm scores increased from 59.5 20.7 to 93.8 7.9 (p &lt; 0.0001). The mean CD index decreased from 1.2 (range 1.0-1.56) to 0.88 (range 0.6-1.0) postoperatively. No patellar redislocations or progression of patellofemoral osteoarthritis were observed. One superficial infection resolved with oral antibiotics, and one tibial tubercle fracture was successfully managed with plate fixation. Four patients had delayed recovery but recovered full function without long-term sequelae.<br />Conclusion. 1. TTO-DM led to excellent functional outcomes and complete prevention of redislocation, even in patients with CD indices ≤1.2. 2. These findings suggest that surgical indications for distalization may be expanded based on trochlear morphology rather than the CD index alone. 3. Although limited by its retrospective design and small sample size, this study provides one of the longest follow-up durations, supporting this approach in patellofemoral instability management.</p>.
To investigate the predictive value of inflammatory, liver, renal, and cardiac biomarkers in relation to the progression of multiorgan failure (MOF) in COVID-19 patients. A primary goal was to identify which of these markers served as the most discriminative predictors of disease severity. A retrospective cohort study was carried out between December 2020 and February 2023 to collect data from 1,803 COVID-19 patients from data records in different hospitals in Makkah, Saudi Arabia. The ROC curve analysis evaluated the diagnostic and predictive performance of selected biomarkers. Inflammatory markers such as CRP and D-dimer were evaluated, but ferritin showed only a moderate discriminatory ability for disease severity with moderate predictive value (AUC 0.621). Liver enzymes, especially AST (aspartate aminotransferase), and ALT (alanine aminotransferase), emerged as strong MOF predictors, with AST achieving an AUC of 1.000, indicating perfect sensitivity and specificity as a predictor. Renal markers like creatinine and BUN showed limited predictive power. Among cardiac biomarkers, LDH (lactate dehydrogenase), showing a robust predictive capability for MOF, demonstrated high predictive capability (AUC 0.969), and CK also performed well. The AST and LDH were identified as the most reliable predictors of MOF in COVID-19 patients. These findings emphasize the importance of biomarker profiling in clinical risk assessment and early intervention. Liver and cardiac markers should be prioritized in evaluating severe COVID-19 cases to support better patient management and outcomes.
Brucellosis is a zoonotic disease that poses a significant threat to public health and the development of animal husbandry, making early and rapid diagnosis essential for effective control. In this study, a double-antigen sandwich time-resolved fluorescent immunochromatographic assay (EuCM-TRFICA) utilizing europium chelate microsphere (EuCM) was developed for the highly sensitive quantification of Brucella antibodies in human serum. This is the first report of a double-antigen sandwich TRFICA utilizing Brucella LPS, which enables the detection of total antibodies with superior sensitivity compared to the previously reported colloidal gold immunochromatographic assay (AuNP-ICA) and time-resolved fluorescence LFIA (TF-LFIA). Under the optimal parameters, EuCM-TRFICA exhibited a satisfactory linear range from 0.156 to 5.00 IU/mL, and its limit of detection (0.145 IU/mL) was lower than that (0.3125 IU/mL) of AuNP-ICA. Furthermore, it effectively identified positive samples missed by the serum agglutination test (SAT) or the rose bengal test (RBT). EuCM-TRFICA exhibited high specificity, with only weak cross-reactivity observed against Yersinia enterocolitica O9, which has been reported to share similar antigenic epitopes. Precision evaluation showed that both intra- and inter-assay coefficients of variation (CV) were below 10.55%. The recoveries of spiked samples ranged from 91.94% to 108.85%, confirming reliable accuracy. In the testing of 49 confirmed positive samples and 60 negative samples, EuCM-TRFICA achieved 100% accuracy, outperforming SAT (92.7%), RBT (86.2%), and ELISA (97.2%). In conclusion, the EuCM-TRFICA exhibits excellent performance in terms of sensitivity, quantitative capability, and detection speed. It features simple operation, a short detection time of 15 min, quantitative readout, and controllable costs. This assay shows promise as a potentially efficient technical tool for early on-site screening and epidemic surveillance in this preliminary cohort, which may be valuable for managing human brucellosis.
Land use change is a key factor regulating water conservation function of river basins. Qiantang River Basin is a crucial ecological barrier and water supply source for the Yangtze River Delta region. Understanding the long-term changes in land use and water conservation function has significant importance for regional sustainable development and water security. We used the InVEST model to analyze the changes in land use patterns and water conservation functions in the Qiantang River Basin in 2000, 2010, and 2020, and used random forest model to analyze the driving factors of water conservation function. The results showed that land use conversion in the Qiantang River Basin during 2000-2020 mainly occurred in the transfer among cropland, forest and construction land. A total area of 1317.26 km2 of cropland and 158.67 km2 of forest had been converted into construction land, while 1301.15 km2 of forest had been transformed into cropland. The total water conservation volume of the Qiantang River Basin in 2000, 2010, and 2020 was 1053090.56×104, 1431102.37×104, and 1204484.25×104 m3 respectively, showing a trend of increasing first and then decreasing. There were significant differences in water conservation capacity among land use types. The total water conservation amount during the study period ranked as forest>cropland>construction land>water area>grassland>shrubland>unused land. Precipitation exerted the greatest impact on water conservation capacity, followed by forest area, cropland area and slope. During the study period, the dominant role of precipitation was gradually weakening, while the regulatory effects of underlying surface factors such as forest area and water area had increased. The results would provide a basis for land use pattern optimization and water resources allocation in the Qiantang River Basin, and offer a reference for restoring water conservation functions in other river basins. 土地利用格局变化是调控流域水源涵养功能的关键因子。钱塘江流域作为长三角地区重要生态屏障和水源供给地,分析其长时间序列土地利用及水源涵养功能的变化特征及其对区域可持续发展和水安全保障具有重要意义。本研究利用InVEST模型分析了钱塘江流域2000、2010和2020年的土地利用格局和水源涵养功能变化情况,并利用随机森林模型分析水源涵养功能的驱动因素。结果表明:2000—2020年,钱塘江流域土地利用类型转化主要集中在耕地、林地与建设用地之间,有1317.26 km2耕地和158.67 km2林地被分别改造成建设用地,同时有1301.15 km2林地转为耕地。钱塘江流域2000、2010和2020年的水源涵养总量分别为1053090.56×104、1431102.37×104和1204484.25×104 m3,呈现先增加后减少的趋势。不同地类的水源涵养量有明显差异,研究期间水源涵养总量依次为林地＞耕地＞建设用地＞水域＞草地＞灌木＞未利用地。降水量对水源涵养能力的影响最大,其次是林地面积、耕地面积和坡度。研究期间,降水量的主导作用在逐渐减弱,而林地面积、水域面积等地表下垫面的调控作用在明显增强。研究结果能为钱塘江流域土地利用格局和水资源配置提供依据,也可为其他流域的水源涵养功能恢复提供借鉴。.
In origins-of-life research, a key challenge is to explain the emergence of polymers of sufficient length to confer the complex functions needed for genetic inheritance. Previous studies have demonstrated that prebiotic environments enabling multilevel selection can facilitate the survival of cooperating polymers such as ribozymes, which allows that complex functions undertaken today by long polymers might have originated from multiple simpler functions undertaken by shorter polymers. To further investigate this possibility, we developed a new computational model of cooperative catalytic and replicating polymer systems that avoids tracking all possible polymer sequences. This approach scales well computationally, avoiding the need to set an a priori cap on polymer length. We first validated this model by replicating key conclusions of previous studies, for example that the persistence of cooperative synthetase-ligase systems is facilitated by both intrinsic factors (shorter length, higher catalytic efficiency) and by factors that promote multilevel-selection (compartmentalization, slower diffusion). We then explored the effects of introducing a mutation inhibitor into a cooperative synthetase-ligase system. The results support the possibility that mutation inhibition could have arisen, not through the appearance of a single proofreading polymerase, but through the emergence of distinct, mutation inhibiting catalysts.
Food contaminated with Salmonella remains a major cause of foodborne illness worldwide, posing a significant public health concern. Rapid detection of viable Salmonella in food is essential for effective food safety management. Viability PCR (vPCR) techniques using photo-reactive dyes such as PMA (Propidium monoazide) or PMAxx have emerged as promising approaches to suppress DNA amplification from membrane-compromised (dead) cells. However, most previous PMA-qPCR studies have mainly evaluated artificially contaminated meat samples, while their performance in naturally contaminated food matrices with complex microbiota remains limited. This study optimized a PMAxx real-time PCR assay for the detection of viable and viable but non-culturable (VBNC) Salmonella and evaluated its application in retail meat samples. The PMAxx real-time PCR technique effectively eliminated signals from 108 CFU/mL of dead cells in culture media samples. The limit of detection (LOD) was 10 CFU/g of viable cells in spiked chicken samples following 18 h of pre-enrichment in Buffered Peptone Water (BPW). Under 0.85% NaCl at 4 °C, Salmonella entered the VBNC state after 32 weeks, while PMAxx real-time PCR still detected signals. The method was evaluated using 33 chicken and pork samples collected from supermarkets and traditional markets. A total of 23 samples tested positive, including 13 samples detected after only 6 h of pre-enrichment. The results show 100% agreement with the traditional culture method performed in accordance with ISO 6579-1:2017. These findings indicate that the PMAxx real-time PCR assay can be applied for the rapid screening of viable Salmonella contamination in fresh food samples.
To study the associations of dietary intake of A and E vitamins, as well as plasma retinols, carotenoids, and tocopherols in relation to development of islet autoimmunity and progression to T1D. The Environmental Determinants of Diabetes in the Young (TEDDY) Study followed 7659 newborns with genetic susceptibility to T1D for 6 years in the USA, Finland, Germany, and Sweden. Dietary vitamin intake was assessed repeatedly with 3-day food-records in full cohort at ages 6 months to 6 years. Plasma retinols, carotenoids, and tocopherols were analysed in a nested case-control setting with 359 children with islet autoimmunity and 1033 matched controls. In the full cohort analyses, dietary intake of retinol, β-carotene, and vitamin E was not associated with the risk of islet autoimmunity or progression to T1D. Further, none of the plasma retinol, carotenoid, and tocopherol biomarkers were associated with islet autoimmunity or T1D in the full nested case-control analyses. We observed effect modification by country, breastfeeding, sex, and follow-up time for both intake and biomarkers of vitamins on the risk of islet autoimmunity or T1D, and some subgroup associations. Finally, a plasma carotenoid metabolite (likely zeinoxanthin) (OR 0.61, 95% CI 0.39, 0.95, p = 0.03) and γ-carotene at 6 months (OR 0.65, 95% CI 0.45, 0.94, p = 0.02) were inversely associated with the odds of developing GADA-first. Retinol, carotenoids and tocopherols were not consistently associated with islet autoimmunity. This study adds to the understanding of factors and their interactions related to T1D development.
Acne is a chronic skin condition that primarily affects adolescents and young adults but can persist into adulthood. It can have repercussions on physical and mental health, self-esteem, and body image. The increasing use of social media for health information and peer support offers an opportunity to explore real-life experiences with acne. This study aims to analyze social media messages from users in the United States and the United Kingdom using artificial intelligence to assess the impact of acne on quality of life (QoL), identify discussion topics, and explore unmet needs. The data were extracted from public platforms using a query containing the word "acne" between January 1 and December 31, 2024. Data cleaning and filtering were performed using natural language processing, machine learning methods, and algorithms. Biterm topic modeling was used to identify the main discussion topics, and QoL impact was assessed using a deep learning algorithm adapted from the EuroQol 5-Dimension Questionnaire or the 36-Item Short Form Health Survey. Unmet needs were identified through manual annotation using the saturation method. A total of 646,809 messages posted by 432,234 users were identified. The main topics included skincare routines and product recommendations (n=154,907, 23.9%), acne scars (n=135,643, 21%), and general treatment information (n=97,177, 15%). Engagement varied across topics and platforms. On Instagram, dietary and nutritional strategies (0.16%, SD 6.36%) showed the highest mean engagement, followed by skincare routines and product recommendations (0.11%, SD 4.81%). In general, engagement scores were higher in the United Kingdom compared to the United States across all topics. On TikTok, content about makeup and acne had the highest mean engagement score (3.03%, SD 92.65%). Overall, 52.9% (228,613/432,234) of the users expressed at least 1 QoL impact, most frequently related to signs and symptoms (175,604/228,613, 76.8%), social functioning (n=149,234, 65.3%), mental health (n=107,155, 46.9%), and cost (n=62,008, 27.1%). Of 3200 annotated messages, 582 contained unmet needs, including effective solutions for hormonal acne (111/582, 19.1%), clarity in identifying acne triggers (n=84, 14.4%), treatment guidance (n=68, 11.7%), and psychological support (n=68, 11.7%). This study revealed the significant physical, psychological, social, and financial impact of acne on QoL and identified several unmet needs. Given the growing role of social media, these findings highlight opportunities for dermatologists and health professionals to educate and engage with the acne community through digital platforms.
The Quality-of-Life Primary Ciliary Dyskinesia (QOL-PCD) instrument is the only health-related QOL instrument specific to measuring patient-reported outcomes in PCD patients. The adaptation and validation of the Spanish version of this questionnaire is a crucial step in formulating effective healthcare strategies that will improve the quality of life of these patients within the Spanish context. In this regard, the objective of the study was to cross-culturally adapt and pilot-validate a Spanish version for patients with PCD. A total of 21 Spanish patients with PCD participated in this study. The QOL-PCD was translated and cross-culturally adapted following BESTCILIA Work Package 4 and ISPOR recommendations, respectively. Pilot psychometric validation was explored through internal consistency (Cronbach's α) and construct validity studies. Stability and convergent validity were studied in eight patients. QoL analyses were performed using Pearson correlations and a Welch t-test. The Spanish QOL-PCD questionnaire included 40 items evaluating the ten original scales. All participants demonstrated a clear understanding of the questionnaire. All scales shown moderate to good internal consistency. Test-retest reliability proved particularly strong for Physical Functioning, Vitality, Emotional Functioning, Role and Social Functioning. SF-36 and SNOT-22 comparisons demonstrated moderate to high correlations. As hypothesized, a positive correlation was found between lower respiratory symptoms and history of lung transplantation. The Spanish QOL-PCD questionnaire has robust psychometric properties and is therefore a validated tool for assessing quality of life (QoL) in Spanish adults with primary ciliary dyskinesia (PCD). This work will improve patient care and inform future PCD research.
The last survivors of the northern white rhinoceros (NWR, Ceratotherium simum cottoni), once widespread across Central and East Africa, are two non-reproductive females under continuous human protection. In order to prevent the extinction of this white rhinoceros subspecies, the BioRescue consortium founded in 2019 has developed an innovative reproductive biotechnology program. This holistic rescue strategy implements (i) advanced assisted reproductive technologies (aART) including in vitro fertilization (IVF) and embryo transfer, (ii) stem cell associated techniques (SCAT) for establishing in vitro gametogenesis (IVG), and (iii) a pangenetic rescue strategy (PRS) that uses the full spectrum of genetic diversity still available by incorporating DNA sequence information from the globally available NWR-museum specimens combined with gene editing to enrich the genetic diversity of the future living population. Key milestones of the BioRescue consortium are: 26 non-surgical oocyte collections in NWR followed by in-vitro oocyte maturation and embryo generation, first pregnancy in a southern white rhino (SWR) surrogate after heterologous embryo transfer, establishment of two SWR embryonic stem cell (ESC) lines, NWR induced pluripotent stem cell (iPSC) lines derived from somatic tissue, and robust primordial germ cell-like cell (PGCLC) induction as a first step towards IVG. Beyond biological and technical challenges, an essential part of the BioRescue project's operational framework addresses the ethical dimension of this new approach in conservation, ensuring transparency, animal welfare, and societal accountability. This multidisciplinary strategy offers a replicable model in conservation science for rescuing critically endangered or practically extinct species, linking the most advanced reproductive technologies with ethical oversight to safeguard biodiversity.
Inoculum preparation of Mycobacterium tuberculosis isolates is a critical step for the reproducibility of drug susceptibility tests (DST) and the prevention of false results. In this study, we investigated the effect of increasing inoculum sizes of M. tuberculosis isolates on DST. For this purpose, primary drug susceptibilities of five ATCC strains and 24 M. tuberculosis isolates were determined by the proportion method on 7H10 agar using six different inoculum sizes (10-2 dilution of McFarland no 1 as reference inoculum and McFarland no 0.5-1-2-3-4). Among the tested isolates (including ATCC strain), 22 of them had DST results at increasing inoculum sizes that were 100% consistent with DST results in the reference inoculum and MGIT-960. For alone streptomycin (STR), in seven isolates tested, DST results at increasing inoculum sizes were 100% consistent with MGIT-960 results but were inconsistent with results determined with the reference inoculum. These isolates were found to be susceptible to STR with reference inoculum, but resistant to STR in MGIT-960 and increasing inoculum sizes. Our study reveals that inoculum size does not affect DST results of M. tuberculosis; on the contrary, increasing the inoculum may have positive results for some antibiotics, such as STR.
Infectious diseases remain a significant global health challenge, necessitating rapid and accurate diagnostic tools to distinguish between bacterial and viral infections. Serum amyloid A (SAA) and C-reactive protein (CRP) are well-established biomarkers of infectious diseases, with CRP typically indicating bacterial infections. This study presents a novel dual-color ultrabright aggregation-induced emission luminogens (AIEgens) nanoparticle-based lateral flow immunoassay (DC-AIENPs-LFIA) for the simultaneous quantitative detection of SAA and CRP. We developed a dual-AIEgens-encapsulated nanoparticle with a large Stokes shift and a high quantum yield of 93% via fluorescence resonance energy transfer, thereby achieving high sensitivity and specificity. The DC-AIENPs-LFIA demonstrated excellent detection performance, with sensitivities of 1.77 μg/mL for SAA and 0.45 μg/mL for CRP, and showed strong correlations with chemiluminescence immunoassay results in clinical serum samples. The assay also exhibited robust discriminatory capacity in differentiating infected from non-infected individuals solely based on SAA and CRP concentrations. Meanwhile, the SAA/CRP ratio showed promise in differentiating bacterial from viral infections, with an area under the receiver operating characteristic curve of 0.95. This study highlights the potential of DC-AIENPs-LFIA as a rapid, sensitive, and reliable point-of-care test for infectious disease diagnosis. It also provides a reference for the development of biosensors capable of multiplex biomarker detection with synergistic diagnostic value.
Hepatitis B virus (HBV) infection poses a significant public health challenge, leading to liver cirrhosis and hepatocellular carcinoma (HCC). The clinical course of HBV infection is influenced by viral, environmental, and host genetic variants. Recently, a Cirrhosis Risk Score (CRS) based on seven genetic variants has been developed to identify individuals at risk of developing cirrhosis. This study aimed to assess the predictive value of seven gene signatures in Egyptian patients with HBV infection as molecular biomarkers for cirrhosis. In this cross-sectional study, 170 HBV-infected patients exhibiting various degrees of liver fibrosis were recruited. Genotyping for the seven gene SNPs was performed using allelic discrimination assays. Subsequently, the non-invasive scores APRI and FIB-4 were computed and compared with CRS values. Furthermore, an in silico analysis was performed to examine the alterations in gene expression associated with the progression of fibrosis. The findings revealed a significantly higher prevalence of high CRS scores in late fibrosis (p = 0.006). The mean of CRS was higher in late than in early fibrosis (0.70 vs. 0.60; P = 0.003). However, APRI and FIB-4 scores did not differ significantly across the groups. The ROC analysis indicated that the CRS score can distinguish between patients with early and late fibrosis, with an AUC of 0.63 (p = 0.0034). The risk genotypes in DEGS1 (rs4290029), STXBP5L (rs17740066), and AQP2 (rs2878771) were associated with late fibrosis. According to the in silico analysis, there is a remarkable upregulation of DEGS1 and a downregulation of STXBP5L in late fibrosis as compared to early fibrosis. While the AQP2 showed no significant variation between early and late fibrosis. The CRS score showed a modest predictive ability in HBV-infected Egyptian patients; it can be used in conjunction with imaging and clinical tools to improve risk stratification. Moreover, DEGS1, STXBP5L, and AQP2 genetic variants demonstrated a significant association with cirrhosis risk. However, only the DEGS1 and STXBP5L genes showed dysregulated expression levels, suggesting their potential relevance as diagnostic biomarkers in liver fibrosis.
Ofloxacin, a fluoroquinolone antibiotic, is a significant contaminant in food products. Its levorotatory form (S-OFL) exhibits a bactericidal activity considerably higher than the dextrorotatory form (R-OFL) and is commonly employed in veterinary medicines. In this work, a highly sensitive immunochromatographic analysis (ICA) method for biologically active S-OFL was developed. A bimetallic core@shell Au@Pd nanozyme was synthesized to serve as the label. Thanks to its peroxidase-like catalytic activity, the nanozyme enhances the color signal on test strips, thereby improving assay sensitivity. The instrumental limit of detection and cutoff values were determined to be 1.4 pg/mL and 0.1 ng/mL, corresponding to 60- and 10-fold improvements compared with conventional gold nanoparticle-based ICA. The cross-reactivity with R-OFL was only 6.3%, demonstrating the high specificity toward target isomer. The ICA was successfully applied to detecting S-OFL in milk, achieving recoveries of 77-99%. The entire nanozyme-enhanced analysis can be completed within 20 min. This method shows potential for monitoring various toxicants in food products.
Single-cell RNA sequencing (scRNA-seq) resolves cell types and molecular phenotypes within heterogeneous specimens but typically requires fresh, high-quality single-cell suspensions processed immediately to preserve transcriptional profiles. This constraint complicates samples with long preparation times and prevents collection at remote sites lacking single-cell instrumentation. Several commercial assays now enable preservation at the point of collection through fixation or cryopreservation, allowing processing to occur months later. The Association of Biomolecular Research Facilities' DNA Sequencing and Genomics and Bioinformatics Research Groups undertook a cross-platform, multisite study to assess the performance and reproducibility of three such platforms: 10x Genomics FLEX, Parse Biosciences Evercode WT v2, and Honeycomb Bio HIVE. Total leukocytes and peripheral blood mononuclear cells (PBMCs) were isolated from a single healthy individual, with EasySep reagent used for red blood cell depletion of the leukocyte fraction. Cells were characterized by a 21-color flow cytometry panel to provide a reference, and the remaining material was fixed or cryopreserved according to each platform's protocol. Preserved leukocyte samples were prepared in parallel by two technicians ("A" and "B" replicates) and distributed to multiple ABRF member core facilities for downstream processing, while fresh leukocytes processed with the 10x 3' v3.1 (3pGEX) chemistry served as a reference. Libraries were sequenced at a central site, and performance was evaluated across standard scRNA-seq quality control metrics, gene and transcript detection sensitivity, cell-type discovery and annotation, differential expression, and correlation analyses. Data from all platforms integrated effectively and produced concordant results for cell-type annotation and relative abundance, with cell-type proportions broadly consistent with the flow cytometry reference. However, platform-specific expression signatures were evident for a subset of genes, and cross-site reproducibility varied between methods, with the FLEX workflow showing greater susceptibility to technical variation introduced during on-site sample processing. Preservation-based methods (FLEX and HIVE) showed better retention of fragile granulocyte populations than fresh samples processed with the 10x 3pGEX workflow. Improvements to preservation methods are changing how single-cell research is conducted by decoupling sample collection from downstream processing. Our investigation into the performance and reproducibility of each platform provides a resource to help investigators and core facilities select the most appropriate single-cell preservation workflow given their sample type, cell populations of interest, sample collection logistics, and laboratory infrastructure constraints.
Currently, the most commonly used methods for detecting Mycoplasma are the culture method and the indicator cell culture method. However, both approaches exhibit low sensitivity and are incapable of detecting low-concentration contamination. In addition, the detection period may extend up to 28 days, which is unsuitable for rapid screening and may delay timely contamination control measures. To address these limitations, a Mycoplasma detection method based on nucleic acid amplification technology (NAT) was developed following a comparative analysis of gene sequences from various Mycoplasma species. The method was validated with respect to its detection performance and its applicability to biological product samples. DNA was extracted from Mycoplasma-contaminated samples using a magnetic bead-based nucleic acid extraction method. Universal primers were designed based on the highly conserved 16S rRNA gene sequence of Mycoplasma, and amplification was performed using multiplex quantitative PCR (qPCR) with fluorescent probes. The limit of detection (LOD) was established based on statistics of 24 replicates. Method specificity and robustness were evaluated according to the guidelines set by the International Council for Harmonisation of Technical Requirements for Pharmaceuticals for Human Use (ICH Q2), while sample applicability was assessed in accordance with the European Pharmacopoeia (EP) <2.6.7>. The NAT-based Mycoplasma detection method enabled rapid, qualitative identification of Mycoplasma contamination. The validated LOD was 10 CFU/mL, and the method met predefined requirements for sensitivity, specificity, and robustness. To assess applicability, real biological product samples, including monoclonal antibodies, antibody fusion proteins, bispecific antibodies, and trispecific antibodies, were spiked with 10 CFU/mL of standard Mycoplasma strains. All spiked samples tested positive. These findings confirm that the NAT-based Mycoplasma detection method is suitable for process control and product release testing in the production of biological products.
The p47ING1a isoform of the ING1 tumor suppressor regulates cellular senescence through Rb-dependent pathways via its plant homeodomain (PHD) zinc-finger, which recognizes the H3K4me3 histone mark. However, the mutational landscape of p47ING1a and the functional consequences of PHD-domain nonsynonymous single-nucleotide polymorphisms (nsSNPs) remain poorly characterized. This study aimed to identify and structurally evaluate the most deleterious nsSNPs in p47ING1a and clarify their potential role in disrupting ING1 tumor-suppressor activity. A total of 347 missense nsSNPs were retrieved from the NCBI dbSNP database and screened using 12 sequence-based computational tools. Variants consistently predicted as deleterious were further evaluated by I-Mutant stability analysis and ConSurf evolutionary conservation profiling. Three-dimensional structural modeling was performed using AlphaFold3, refined through GalaxyRefine, and validated by ERRAT, PROCHECK, and TM-align. Mutation-induced structural and binding effects were assessed using Missense3D, mCSM, and BeAtMuSiC. Post-translational modification sites were predicted via NetPhos 3.1, GPS 3.0, BDM-PUB, and NetOGlyc 4.0. Protein-protein interaction networks were constructed using STRING and Gene MANIA. Pan-cancer expression was analyzed through UALCAN and the Human Protein Atlas. Twelve computational tools converged on six high-priority variants, namely, C358S, C374G, W378G, F379V, S382L, and R400P. All localized exclusively within the PHD zinc-finger domain, residues 353-402. All six mutations were consistently predicted to destabilize the p47ING1a protein across multiple stability analyses. Six nsSNPs in the PHD domain of p47ING1a are predicted to disrupt protein stability, H3K4me3 binding, and Sin3A/HDAC complex interactions, thereby impairing ING1 tumor-suppressor function. These findings provide a computational basis for prioritizing variants for experimental validation through site-directed mutagenesis, chromatin-binding assays, and structure-guided therapeutic targeting of the PHD-H3K4me3 interface.
Background: Lung adenocarcinoma (LUAD) is the most common subtype of non-small cell lung cancer (NSCLC) and remains a leading cause of cancer-related mortality worldwide. Aberrant glycosylation contributes to tumor progression by regulating receptor signalling, immune evasion, and metastatic. However, the prognostic and therapeutic relevance of glycosylation-related genes (GRGs) in LUAD has not been comprehensively defined. Therefore, this study aimed to comprehensively evaluate GRG-associated molecular subtypes and their clinical and therapeutic relevance in LUAD. Methods: GRGs were curated from multiple public databases and integrated with transcriptomic and clinical data from The Cancer Genome Atlas LUAD cohort (TCGA-LUAD) with validation in Gene Expression Omnibus (GEO) datasets. Consensus clustering, pathway enrichment, and immune profiling were used to identify glycosylation-associated subtypes. A glycosylation activity scoring (Glyco. marker) was developed to quantify glycosylation features. Drug response prediction was analyzed using OncoPredict and the Genomics of Drug Sensitivity in Cancer (GDSC) database. Single-cell RNA sequencing (scRNA-seq) was analyzed to evaluate cell-type-specific GRG expression. Selected proteins were by immunohistochemistry (IHC) in LUAD tissue microarrays. Results: GRG expression stratified 513 LUAD patients into four molecular clusters with distinct clinical and immune characteristics. The Glyco.High group showed elevated expression of MGAT5 (mannosyl (α-1,6)-glycoprotein β-1,6-N-acetylglucosaminyltransferase), ST6GAL1 (β-galactoside α-2,6-sialyltransferase 1), GALNT7 (polypeptide N-acetylgalactosaminyltransferase 7), and FUT8 (fucosyltransferase 8), frequent tumor protein p53 (TP53) mutations, increased immune checkpoint expression, and enrichment of regulatory T cells. The Glyco. marker score predicted overall survival and was associated with stemness signatures. Drug response prediction suggested reduced sensitivity to platinum chemotherapy and epidermal growth factor receptor (EGFR) inhibitors but increased sensitivity to phosphoinositide 3-kinase/protein kinase B/mechanistic target of rapamycin (PI3K/AKT/mTOR) inhibitors. Conclusion: GRG-based molecular stratification identifies clinically distinct LUAD subtypes associated with immune regulation, tumor stemness, and therapeutic response. The Glyco. marker system provides a potential framework for prognostic assessment and precision oncology strategies in LUAD.