A Post-Marketing Surveillance Study of Nusinersen for Spinal Muscular Atrophy in Routine Medical Practice in China: Interim Results.

INTRODUCTION: Spinal muscular atrophy (SMA) is a rare, autosomal recessive, neuromuscular disease that leads to progressive muscular weakness and atrophy. Nusinersen, an antisense oligonucleotide, was approved for SMA in China in February 2019. We report interim results from a post-marketing surveillance phase 4 study, PANDA (NCT04419233), that collects data on the safety, efficacy, and pharmacokinetics of nusinersen in children with SMA in routine clinical practice in China. METHODS: Participants enrolled in PANDA will be observed for 2 years following nusinersen treatment initiation. The primary endpoint is the incidence of adverse events (AEs)/serious AEs (SAEs) during the treatment period. Efficacy assessments include World Health Organization (WHO) Motor Milestones assessment, the Hammersmith Infant Neurological Examination (HINE), and ventilation support. Plasma and cerebrospinal fluid (CSF) concentrations of nusinersen are measured at each dose visit. RESULTS: Fifty participants were enrolled as of the January 4, 2023, data cutoff: 10 with infantile-onset (≤ 6 months) and 40 with later-onset (> 6 months) SMA. All 50 participants have received at least one dose of nusinersen; 6 have completed the study. AEs were experienced by 45 (90%) participants and were mostly mild/moderate; no AEs led to nusinersen discontinuation or study withdrawal. Eleven participants experienced SAEs, most commonly pneumonia (n = 9); none were considered related to study treatment. Stability or gain of WHO motor milestone was observed and mean HINE-2 scores improved in both subgroups throughout the study. No serious respiratory events occurred, and no permanent ventilation support was initiated during the study. Pre-dose nusinersen CSF concentrations increased steadily through the loading-dose period, with no accumulation in plasma after multiple doses. CONCLUSION: Nusinersen was generally well tolerated with an acceptable overall safety profile, consistent with the known safety of nusinersen. Efficacy, safety, and nusinersen exposure are consistent with prior observations. These results support continuing PANDA and evaluation of nusinersen in Chinese participants with SMA. TRIAL REGISTRATION: ClinicalTrials.gov identifier, NCT04419233.

Blocking SLC7A11 attenuates the proliferation of esophageal squamous cell carcinoma cells.

The role of ferroptosis-associated gene SLC7A11 in esophageal cancer progression is largely unknown, therefore, the effects of blocking SLC7A11 on esophageal squamous cell carcinoma (ESCC) cells are evaluated. Results showed that SLC7A11 was overexpressed in ESCC tissues both in mRNA and protein levels. Blocking SLC7A11 using Erastin suppressed the proliferation and colony formation of ESCC cells, decreased cellular ATP levels, and improved ROS production. Sixty-three SLC7A11-binding proteins were identified using the IP-MS method, and these proteins were enriched in four signaling pathways, including spliceosome, ribosome, huntington disease, and diabetic cardiomyopathy. The deubiquitinase inhibitors PR-619, GRL0617, and P 22077 could reduce at least 40% protein expression level of SLC7A11 in ESCC cells, and PR-619 and GRL0617 exhibited suppressive effects on the cell viability and colony formation ability of KYSE30 cells, respectively. Erastin downregulated GPX4 and DHODH and also reduced the levels of ß-catenin, p-STAT3, and IL-6 in ESCC cells. In conclusion, SLC7A11 was overexpressed in ESCC, and blocking SLC7A11 using Erastin mitigated malignant phenotypes of ESCC cells and downregulated key ferroptosis-associated molecules GPX4 and DHODH. The therapeutic potential of targeting SLC7A11 should be further evaluated in the future.

Crotonoids A-H, Labdane-Type Diterpenoids with Anti-Inflammatory Activity from Croton sublyratus.

Croton sublyratus (Euphorbiaceae) is a traditional medicinal plant used by the Thai populace to treat helminthic infections and dermatologic conditions. In present study, eight new labdane-type diterpenoids, crotonoids A-H (1-8) and one known analogue (9) were isolated from the aerial parts of C. sublyratus. Compounds 6 and 7 belong to the rare class of 14,15-dinor-labdane diterpenoids. Compound 8 exhibited a rare 14,15,17-trinor-labdane skeleton. The structures of all these diterpenoids were elucidated by spectroscopic data analysis, electronic circular dichroism calculations, and single-crystal X-ray diffraction analysis. Compound 9 exhibited moderate anti-inflammatory activity via the inhibition of NO production in lipopolysaccharide-induced RAW 264.7 cells.

Lauinoids A-X: Labdane-type diterpenoids with anti-inflammatory activity from Croton laui.

Croton laui (Euphorbiaceae) is a traditional medicinal plant used by the Li ethnic group in China to treat headaches, stomachaches, and diphtheria. To understand the pharmacological basis of its medicinal use, an extensive investigation of the ethanolic extract of the bark of C. laui was performed. After repeated chromatography, twenty-four undescribed labdane-type diterpenoids, lauinoids A-X (1-24), and five known analogs (25-29) were isolated. Their structures and absolute configurations were established using a combination of spectroscopic analyses, electronic circular dichroism, nuclear magnetic resonance calculations, and single-crystal X-ray diffraction. Among them, compounds 1-3 exhibited an 11(12 â†’ 13)-abeo-16-nor-labdane skeleton, which originated putatively from 9 through a plausible pathway that involves a semipinacol rearrangement process. Compounds 11 and 12 belong to the rare class of 14,15-dinor-labdane diterpenoids. Compounds 18 and 28 exhibited substantial inhibitory effects by suppressing lipopolysaccharide-induced NO production in RAW 264.7 macrophages, with IC50 values of 3.37 ± 0.23 and 5.82 ± 0.28 µM, respectively. This study has greatly expanded the chemical diversity of labdane diterpenoids from C. laui and will guide future research on this ethnomedicinal plant.

Oral microbiota dysbiosis alters chronic restraint stress-induced depression-like behaviors by modulating host metabolism.

Studies have shown that the microbiota-gut-brain axis is highly correlated with the pathogenesis of depression in humans. However, whether independent oral microbiome that do not depend on gut microbes could affect the progression of depression in human beings remains unclear, neither does the presence and underlying mechanisms of the microbiota-oral-brain axis in the development of the condition. Hence this study that encompasses clinical and animal experiments aims at investigating the correlation between oral microbiota and the onset of depression via mediating the microbiota-oral-brain axis. We compared the oral microbial compositions and metabolomes of 87 patients with depressive symptoms versus 70 healthy controls. We found that the oral microbial and metabolic signatures were significantly different between the two groups. Significantly, germ-free (GF) mice transplanted with saliva from mice exposing to chronic restraint stress (CRS) displayed depression-like behavior and oral microbial dysbiosis. This was characterized by a significant differential abundance of bacterial species, including the enrichment of Pseudomonas, Pasteurellaceae, and Muribacter, as well as the depletion of Streptococcus. Metabolomic analysis showed the alternation of metabolites in the plasma of CRS-exposed GF mice, especially Eicosapentaenoic Acid. Furthermore, oral and gut barrier dysfunction caused by CRS-induced oral microbiota dysbiosis may be associated with increased blood-brain barrier permeability. Pseudomonas aeruginosa supplementation exacerbated depression-like behavior, while Eicosapentaenoic Acid treatment conferred protection against depression-like states in mice. These results suggest that oral microbiome and metabolic function dysbiosis may be relevant to the pathogenesis and pathophysiology of depression. The proposed microbiota-oral-brain axis provides a new way and targets for us to study the pathogenesis of depression.

Massively parallel in vivo Perturb-seq reveals cell-type-specific transcriptional networks in cortical development.

Leveraging AAVs' versatile tropism and labeling capacity, we expanded the scale of in vivo CRISPR screening with single-cell transcriptomic phenotyping across embryonic to adult brains and peripheral nervous systems. Through extensive tests of 86 vectors across AAV serotypes combined with a transposon system, we substantially amplified labeling efficacy and accelerated in vivo gene delivery from weeks to days. Our proof-of-principle in utero screen identified the pleiotropic effects of Foxg1, highlighting its tight regulation of distinct networks essential for cell fate specification of Layer 6 corticothalamic neurons. Notably, our platform can label >6% of cerebral cells, surpassing the current state-of-the-art efficacy at <0.1% by lentivirus, to achieve analysis of over 30,000 cells in one experiment and enable massively parallel in vivo Perturb-seq. Compatible with various phenotypic measurements (single-cell or spatial multi-omics), it presents a flexible approach to interrogate gene function across cell types in vivo, translating gene variants to their causal function.

Enhanced external counterpulsation treatment improves multi-organ hemodynamics for postoperative liver transplantation patient. A case report.

INTRODUCTION: Post liver transplantation (LT) patients endure high morbidity rate of multi-organ ischemic symptoms following reperfusion. We hypothesize that enhanced external counterpulsation (EECP) as a typical non-invasive assisted circulation procedure, which can efficiently inhibit the relative ischemic symptoms via the systemic improvement of hemodynamics. CASE PRESENTATION: A 51-year-old male patient, 76 kg, 172 cm, received orthotopic LT surgery for viral hepatitis B induced acute-on-chronic liver failure hepatic failure. His medical records revealed ischemic symptoms in multi-organ at the time of hospital discharge, including headache, refractory insomnia, abdominal paralysis, and lower limb pain. The EECP treatment was introduced for assisted rehabilitation and to improve the postoperative quality of life. Doppler Ultrasound examination showed significant augmentation of blood flow volume in the carotid arteries, the hepatic artery, the portal vein and the femoral artery during EECP intervention. A standard 35-hour EECP treatment led to significant improvement in quality of life, e.g. sleep quality and walking ability. CONCLUSION: We report a case of multi-organ ischemic symptoms in a post LT patient. EECP treatment can significantly improve the quality of life via the systematic promotion of hemodynamics.

Investigating the Impact of Gut Microbiota on Gout Through Mendelian Randomization.

Background: The relationship between gout and gut microbiota has attracted significant attention in current research. However, due to the diverse range of gut microbiota, the specific causal effect on gout remains unclear. This study utilizes Mendelian randomization (MR) to investigate the causal relationship between gut microbiota and gout, aiming to elucidate the underlying mechanism of microbiome-mediated gout and provide valuable guidance for clinical prevention and treatment. Materials and Methods: The largest genome-wide association study meta-analysis conducted by the MiBioGen Consortium (n=18,340) was utilized to perform a two-sample Mendelian randomization investigation on aggregate statistics of intestinal microbiota. Summary statistics for gout were utilized from the data released by EBI. Various methods, including inverse variance weighted, weighted median, weighted model, MR-Egger, and Simple-mode, were employed to assess the causal relationship between gut microbiota and gout. Reverse Mendelian randomization analysis revealed a causal association between bacteria and gout in forward Mendelian randomization analysis. Cochran's Q statistic was used to quantify instrumental variable heterogeneity. Results: The inverse variance weighted estimation revealed that Rikenellaceae exhibited a slight protective effect on gout, while the presence of Ruminococcaceae UCG_011 is associated with a marginal increase in the risk of gout. According to the reverse Mendelian Randomization results, no significant causal relationship between gout and gut microbiota was observed. No significant heterogeneity of instrumental variables or level pleiotropy was detected. Conclusion: Our MR analysis revealed a potential causal relationship between the development of gout and specific gut microbiota; however, the causal effect was not robust, and further research is warranted to elucidate its underlying mechanism in gout development. Considering the significant association between diet, gut microbiota, and gout, these findings undoubtedly shed light on the mechanisms of microbiota-mediated gout and provide new insights for translational research on managing and standardizing treatment for this condition.

Stepwise Synthesis of Low-Symmetry Hexacationic Pyridinium Organic Cages.

An efficient approach was developed for the synthesis of the well-known BlueCage by pre-bridging two 2,4,6-tris(4-pyridyl)-1,3,5-triazine (TPT) panels with one linker followed by cage formation in a much improved yield and shortened reaction time. Such a stepwise methodology was further applied to synthesize three new pyridinium organic cages, C2, C3, and C4, where the low-symmetry cages C3 and C4 with angled panels demonstrated better recognition properties toward 1,1'-bi-2-naphthol (BINOL) than the high-symmetry analogue C2 featuring parallel platforms.

Targeting selective autophagy and beyond: From underlying mechanisms to potential therapies.

BACKGROUND: Autophagy is an evolutionarily conserved turnover process for intracellular substances in eukaryotes, relying on lysosomal (in animals) or vacuolar (in yeast and plants) mechanisms. In the past two decades, emerging evidence suggests that, under specific conditions, autophagy can target particular macromolecules or organelles for degradation, a process termed selective autophagy. Recently, accumulating studies have demonstrated that the abnormality of selective autophagy is closely associated with the occurrence and progression of many human diseases, including neurodegenerative diseases, cancers, metabolic diseases, and cardiovascular diseases. AIM OF REVIEW: This review aims at systematically and comprehensively introducing selective autophagy and its role in various diseases, while unravelling the molecular mechanisms of selective autophagy. By providing a theoretical basis for the development of related small-molecule drugs as well as treating related human diseases, this review seeks to contribute to the understanding of selective autophagy and its therapeutic potential. KEY SCIENTIFIC CONCEPTS OF REVIEW: In this review, we systematically introduce and dissect the major categories of selective autophagy that have been discovered. We also focus on recent advances in understanding the molecular mechanisms underlying both classical and non-classical selective autophagy. Moreover, the current situation of small-molecule drugs targeting different types of selective autophagy is further summarized, providing valuable insights into the discovery of more candidate small-molecule drugs targeting selective autophagy in the future. On the other hand, we also reveal clinically relevant implementations that are potentially related to selective autophagy, such as predictive approaches and treatments tailored to individual patients.

Comparison of interventions for Barrett's esophagus: A network meta-analysis.

BACKGROUND AND OBJECTIVE: Barrett's esophagus (BE) is a precancerous condition that has the potential to develop into esophageal cancer (EC). Currently, there is a wide range of management options available for individuals at different pathological stages in Barrett's esophagus (BE). However, there is currently a lack of knowledge regarding their comparative efficacy. To address this gap, we conducted a network meta-analysis of published randomized controlled trials to examine the comparative effectiveness of all regimens. METHODS: Data extracted from eligible randomized controlled trials were utilized in a Bayesian network meta-analysis to examine the relative effectiveness of BE's treatment regimens and determine their ranking in terms of efficacy. The ranking probability for each regimen was assessed using the surfaces under cumulative ranking values. The outcomes under investigation were complete ablation of BE, neoplastic progression of BE, and complete eradication of dysplasia. RESULTS: We identified twenty-three RCT studies with a total of 1675 participants, and ten different interventions. Regarding complete ablation of non-dysplastic BE, the comparative effectiveness ranking indicated that argon plasma coagulation (APC) was the most effective regimen, with the highest SUCRA value, while surveillance and PPI/H2RA were found to be the least efficacious regimens. For complete ablation of BE with low-grade dysplasia, high-grade dysplasia, or esophageal cancer, photodynamic therapy (PDT) had the highest SUCRA value of 94.1%, indicating it as the best regimen. Additionally, for complete eradication of dysplasia, SUCRA plots showed a trend in ranking PDT as the highest with a SUCRA value of 91.2%. Finally, for neoplastic progression, radiofrequency ablation (RFA) and surgery were found to perform significantly better than surveillance. The risk of bias assessment revealed that 6 studies had an overall high risk of bias. However, meta-regression with risk of bias as a covariate did not indicate any influence on the model. In terms of the Confidence in Network Meta-Analysis evaluation, a high level of confidence was found for all treatment comparisons. CONCLUSION: Endoscopic surveillance alone or PPI/H2RA alone may not be sufficient for managing BE, even in cases of non-dysplastic BE. However, APC has shown excellent efficacy in treating non-dysplastic BE. For cases of BE with low-grade dysplasia, high-grade dysplasia, or esophageal cancer, PDT may be the optimal intervention as it can induce regression of BE metaplasia and prevent future progression of BE to dysplasia and EC.

OsPRMT6a-mediated arginine methylation of OsJAZ1 regulates jasmonate signaling and spikelet development in rice.

Although both protein arginine methylation (PRMT) and jasmonate (JA) signaling are crucial for regulating plant development, the relationship between these processes in spikelet development control remains unclear. Here, we utilized CRISPR/Cas9 technology to generate two OsPRMT6a loss-of-function mutants exhibiting various abnormal spikelet structures. Additionally, we found that OsPRMT6a could methylate arginine residues in the JA signal repressors OsJAZ1 and OsJAZ7. Arginine methylation of OsJAZ1 increased the affinity of OsJAZ1 for the JA receptors OsCOI1a and OsCOI1b in the presence of jasmonates (JAs), subsequently promoting the ubiquitination of OsJAZ1 by the SCFOsCOI1a/OsCOI1b complex and degradation via the 26S proteasome. This process ultimately released OsMYC2, a core transcriptional regulator in the JA signaling pathway, to activate or repress JA-responsive genes, thereby maintaining normal plant (spikelet) development. However, in the osprmt6a-1 mutant, reduced arginine methylation of OsJAZ1 impaired the interaction between OsJAZ1 and OsCOI1a/OsCOI1b in the presence of JAs. As a result, OsJAZ1 proteins became more stable, repressing JA responses, thus causing the formation of abnormal spikelet structures. Moreover, we discovered that JA signaling reduced the OsPRMT6a mRNA level in an OsMYC2-dependent manner, thereby establishing a negative feedback loop to balance JA signaling. Furthermore, we found that OsPRMT6a-mediated arginine methylation of OsJAZ1 likely serves as a switch to tune JA signaling to maintain normal spikelet development under harsh environmental conditions such as high temperatures. Thus, our study established a direct molecular link between arginine methylation and the JA signaling pathway.

CD22-targeted glyco-engineered natural killer cells offer a further treatment option for B-cell acute lymphoblastic leukemia.

Not available.

Ophiocordyceps sinensis preparations combined with the renin-angiotensin system inhibitor for diabetic kidney disease treatment: an umbrella review of systematic reviews and network meta-analysis.

Aims: This study aimed to synthesize the evidence of the comparative effectiveness and safety of Ophiocordyceps sinensis (OS) preparations combined with renin-angiotensin system inhibitors (RASi) for diabetic kidney disease (DKD). Methods: Eight databases were searched from their inception to May 2023. Systematic reviews (SRs) of OS preparations combined with RASi for DKD were identified. Randomized controlled trials (RCTs) from the included SRs and additional searching were performed for data pooling. Cochrane risk-of-bias 2 (RoB 2) tool and AMSTAR 2 were used to evaluate the methodological quality of RCTs and SRs, respectively. A Bayesian network meta-analysis was performed to compare the add-on effect and safety of OS preparations for DKD. The certainty of evidence was graded using the Grading of Recommendations, Assessment, Development, and Evaluation (GRADE) approach. Results: Fourteen SRs were included, whose methodological quality was assessed as high (1/14) or critically low (13/14). After combining additional searching, 157 RCTs were included, involving 13,143 participants. The quality of the RCTs showed some concerns (155/157) or high risk (2/157). Jinshuibao capsules and tablets, Bailing capsules and tablets, and Zhiling capsules were evaluated. Compared to RASi, adding either of the OS capsular preparations resulted in a decreased 24-h urinary total protein levels. OS preparations ranked differently in each outcome. Jinshuibao capsules plus RASi were beneficial in reducing urinary protein, serum creatinine, serum urea nitrogen, and blood glucose levels, with moderate-certainty evidence. No serious adverse events were observed after adding OS to RASi. Conclusion: Combining OS capsular preparations with RASi appeared to be associated with decreased urinary total protein levels in DKD patients. Further high-quality studies are needed to confirm. Systematic Review Registration: INPASY202350066.

Analysis of flow cytometry data from ultrasound-guided lymph node biopsies with two types of needles.

BACKGROUND: In this study, we combined two techniques, ultrasound-guided needle biopsy and flow cytometry (FCM), to explore their value in patients with enlarged lymph nodes. METHODS: We compared the results of 198 needle biopsies on FCM and pathology. Forty-two were done by (fine needle aspiration, FNA), and the remaining 156 with (core needle biopsy, CNB), in 36 of 156 patients, a FNA was performed in the same lymph node after completion of the CNB. Except for five types of pathological entities, the rest were differentiated only detected or undetected tumours as the outcome distinction. RESULTS: Among the 198 needle biopsies, 13 were inadequate specimens, while the remaining 185 had pathological findings, including 47 benign and 138 neoplastic findings. Thirty-six patients underwent puncture with both FNA and CNB, both needles produced identical results by FCM, but more cells were obtained by FNA. Among the pathologically positive results, there were 23 missed diagnoses in FCM, in contrast, evidence of tumours was observed in the FCM images of 15 needle biopsies that reported benign or findings that were inconsistent with pathology, and the final diagnosis was consistent with the FCM in 10 cases. FCM detected haematolymphoid tumours with a sensitivity of 87.8% and a specificity of 91.9%. CONCLUSION: The combination of FCM and ultrasound-guided lymph node needle biopsy can quickly provide guidance for clinical decision-making. We recommend that all lymph node needle biopsies be sent for FCM, the specimen can be obtained by the last puncture with FNA.

Intertumoral Heterogeneity Based on MRI Radiomic Features Estimates Recurrence in Hepatocellular Carcinoma.

BACKGROUND: Hepatocellular carcinoma (HCC) heterogeneity impacts prognosis, and imaging is a potential indicator. PURPOSE: To characterize HCC image subtypes in MRI and correlate subtypes with recurrence. STUDY TYPE: Retrospective. POPULATION: A total of 440 patients (training cohort = 213, internal test cohort = 140, external test cohort = 87) from three centers. FIELD STRENGTH/SEQUENCE: 1.5-T/3.0-T, fast/turbo spin-echo T2-weighted, spin-echo echo-planar diffusion-weighted, contrast-enhanced three-dimensional gradient-recalled-echo T1-weighted with extracellular agents (Gd-DTPA, Gd-DTPA-BMA, and Gd-BOPTA). ASSESSMENT: Three-dimensional volume-of-interest of HCC was contoured on portal venous phase, then coregistered with precontrast and late arterial phases. Subtypes were identified using non-negative matrix factorization by analyzing radiomics features from volume-of-interests, and correlated with recurrence. Clinical (demographic and laboratory data), pathological, and radiologic features were compared across subtypes. Among clinical, radiologic features and subtypes, variables with variance inflation factor above 10 were excluded. Variables (P < 0.10) in univariate Cox regression were included in stepwise multivariate analysis. Three recurrence estimation models were built: clinical-radiologic model, subtype model, hybrid model integrating clinical-radiologic characteristics, and subtypes. STATISTICAL TESTS: Mann-Whitney U test, Kruskal-Wallis H test, chi-square test, Fisher's exact test, Kaplan-Meier curves, log-rank test, concordance index (C-index). Significance level: P < 0.05. RESULTS: Two subtypes were identified across three cohorts (subtype 1:subtype 2 of 86:127, 60:80, and 36:51, respectively). Subtype 1 showed higher microvascular invasion (MVI)-positive rates (53%-57% vs. 26%-31%), and worse recurrence-free survival. Hazard ratio (HR) for the subtype is 6.10 in subtype model. Clinical-radiologic model included alpha-fetoprotein (HR: 3.01), macrovascular invasion (HR: 2.32), nonsmooth tumor margin (HR: 1.81), rim enhancement (HR: 3.13), and intratumoral artery (HR: 2.21). Hybrid model included alpha-fetoprotein (HR: 2.70), nonsmooth tumor margin (HR: 1.51), rim enhancement (HR: 3.25), and subtypes (HR: 5.34). Subtype model was comparable to clinical-radiologic model (C-index: 0.71-0.73 vs. 0.71-0.73), but hybrid model outperformed both (C-index: 0.77-0.79). CONCLUSION: MRI radiomics-based clustering identified two HCC subtypes with distinct MVI status and recurrence-free survival. Hybrid model showed superior capability to estimate recurrence. LEVEL OF EVIDENCE: 3 TECHNICAL EFFICACY STAGE: 2.

Research on high sensitivity piezoresistive sensor based on structural design.

With the popularity of smart terminals, wearable electronic devices have shown great market prospects, especially high-sensitivity pressure sensors, which can monitor micro-stimuli and high-precision dynamic external stimuli, and will have an important impact on future functional development. Compressible flexible sensors have attracted wide attention due to their simple sensing mechanism and the advantages of light weight and convenience. Sensors with high sensitivity are very sensitive to pressure and can detect resistance/current changes under pressure, which has been widely studied. On this basis, this review focuses on analyzing the performance impact of device structure design strategies on high sensitivity pressure sensors. The design of structures can be divided into interface microstructures and three-dimensional framework structures. The preparation methods of various structures are introduced in detail, and the current research status and future development challenges are summarized.

Effects of blood-processing protocols on cell-free DNA fragmentomics in plasma: Comparisons of one- and two-step centrifugations.

BACKGROUND: Cell-free DNA (cfDNA) fragmentomic characteristics are promising analytes with abundant physiological signals for non-invasive disease diagnosis and monitoring. Previous studies on plasma cfDNA fragmentomics commonly employed a two-step centrifugation process for removing cell debris, involving a low-speed centrifugation followed by a high-speed centrifugation. However, the effects of centrifugation conditions on the analysis of cfDNA fragmentome remain uncertain. METHODS: We collected blood samples from 10 healthy individuals and divided each sample into two aliquots for plasma preparation with one- and two-step centrifugation processes. We performed whole genome sequencing (WGS) of the plasma cfDNA in the two groups and comprehensively compared the cfDNA fragmentomic features. Additionally, we reanalyzed the fragmentomic features of cfDNA from 16 healthy individuals and 16 COVID-19 patients, processed through one- and two-step centrifugation in our previous study, to investigate the impact of centrifugation on disease signals. RESULTS: Our results showed that there were no significant differences observed in the characteristics of nuclear cfDNA, including size, motif diversity score (MDS) of end motifs, and genome distribution, between plasma samples treated with one- and two-step centrifugation. The cfDNA size shortening in COVID-19 patients was observed in plasma samples with one- and two-step centrifugation methods. However, we observed a significantly higher relative abundance and longer size of cell-free mitochondrial DNA (mtDNA) in the one-step samples compared to the two-step samples. This difference in mtDNA caused by the one- and two-step centrifugation methods surpasses the pathological difference between COVID-19 patients and healthy individuals. CONCLUSIONS: Our findings indicate that one-step low-speed centrifugation is a simple and potentially suitable method for analyzing nuclear cfDNA fragmentation characteristics. These results offer valuable guidance for cfDNA research in various clinical scenarios.

PI3K/HSCB axis facilitates FOG1 nuclear translocation to promote erythropoiesis and megakaryopoiesis.

Erythropoiesis and megakaryopoiesis are stringently regulated by signaling pathways. However, the precise molecular mechanisms through which signaling pathways regulate key transcription factors controlling erythropoiesis and megakaryopoiesis remain partially understood. Herein, we identified heat shock cognate B (HSCB), which is well known for its iron-sulfur cluster delivery function, as an indispensable protein for friend of GATA 1 (FOG1) nuclear translocation during erythropoiesis of K562 human erythroleukemia cells and cord-blood-derived human CD34+CD90+hematopoietic stem cells (HSCs), as well as during megakaryopoiesis of the CD34+CD90+HSCs. Mechanistically, HSCB could be phosphorylated by phosphoinositol-3-kinase (PI3K) to bind with and mediate the proteasomal degradation of transforming acidic coiled-coil containing protein 3 (TACC3), which otherwise detained FOG1 in the cytoplasm, thereby facilitating FOG1 nuclear translocation. Given that PI3K is activated during both erythropoiesis and megakaryopoiesis, and that FOG1 is a key transcription factor for these processes, our findings elucidate an important, previously unrecognized iron-sulfur cluster delivery independent function of HSCB in erythropoiesis and megakaryopoiesis.