Adaptive functions of structural variants in human brain development.

Quantifying the structural variants (SVs) in nonhuman primates could provide a niche to clarify the genetic backgrounds underlying human-specific traits, but such resource is largely lacking. Here, we report an accurate SV map in a population of 562 rhesus macaques, verified by in-house benchmarks of eight macaque genomes with long-read sequencing and another one with genome assembly. This map indicates stronger selective constrains on inversions at regulatory regions, suggesting a strategy for prioritizing them with the most important functions. Accordingly, we identified 75 human-specific inversions and prioritized them. The top-ranked inversions have substantially shaped the human transcriptome, through their dual effects of reconfiguring the ancestral genomic architecture and introducing regional mutation hotspots at the inverted regions. As a proof of concept, we linked APCDD1, located on one of these inversions and down-regulated specifically in humans, to neuronal maturation and cognitive ability. We thus highlight inversions in shaping the human uniqueness in brain development.

The Histone Variant H3.3 Is Required for Plant Growth and Fertility in Arabidopsis.

Histones are the core components of the eukaryote chromosome, and have been implicated in transcriptional gene regulation. There are three major isoforms of histone H3 in Arabidopsis. Studies have shown that the H3.3 variant is pivotal in modulating nucleosome structure and gene transcription. However, the function of H3.3 during development remains to be further investigated in plants. In this study, we disrupted all three H3.3 genes in Arabidopsis. Two triple mutants, h3.3cr-4 and h3.3cr-5, were created by the CRISPR/Cas9 system. The mutant plants displayed smaller rosettes and decreased fertility. The stunted growth of h3.3cr-4 may result from reduced expression of cell cycle regulators. The shorter stamen filaments, but not the fertile ability of the gametophytes, resulted in reduced fertility of h3.3cr-4. The transcriptome analysis suggested that the reduced filament elongation of h3.3cr-4 was probably caused by the ectopic expression of several JASMONATE-ZIM DOMAIN (JAZ) genes, which are the key repressors of the signaling pathway of the phytohormone jasmonic acid (JA). These observations suggest that the histone variant H3.3 promotes plant growth, including rosette growth and filament elongation.

Large tandem duplications in cancer result from transcription and DNA replication collisions.

Despite the abundance of somatic structural variations (SVs) in cancer, the underlying molecular mechanisms of their formation remain unclear. Here, we use 6,193 whole-genome sequenced tumors to study the contributions of transcription and DNA replication collisions to genome instability. After deconvoluting robust SV signatures in three independent pan-cancer cohorts, we detect transcription-dependent replicated-strand bias, the expected footprint of transcription-replication collision (TRC), in large tandem duplications (TDs). Large TDs are abundant in female-enriched, upper gastrointestinal tract and prostate cancers. They are associated with poor patient survival and mutations in TP53, CDK12, and SPOP. Upon inactivating CDK12, cells display significantly more TRCs, R-loops, and large TDs. Inhibition of G2/M checkpoint proteins, such as WEE1, CHK1, and ATR, selectively inhibits the growth of cells deficient in CDK12. Our data suggest that large TDs in cancer form due to TRCs, and their presence can be used as a biomarker for prognosis and treatment.

[A comparative study of the effects of citrate and heparin anticoagulation on coagulation function and efficacy in children with septic shock undergoing continuous blood purification].

OBJECTIVE: To compare the effects of citrate and heparin anticoagulation on coagulation function and efficacy in children with septic shock undergoing continuous blood purification (CBP), and to provide guidance for CBP anticoagulation in children with septic shock. METHODS: A case control study was conducted. Thirty-seven children with septic shock admitted to the pediatric intensive care unit (PICU) of the First Affiliated Hospital of Gannan Medical University from July 2019 to September 2022 were enrolled as the research subjects. The patients were divided into citrate local anticoagulation group and heparin systemic anticoagulation group according to different anticoagulation methods. The baseline data, the level of coagulation indicators [prothrombin time (PT), activated partial thromboplastin time (APTT), thrombin time (TT), fibrinogen (Fib), D-dimer] before treatment and 1 day after weaning from CBP, serum inflammatory mediators [interleukin-6 (IL-6), tumor necrosis factor-α (TNF-α), hypersensitivity C-reactive protein (hs-CRP), procalcitonin (PCT)], bleeding complications during CBP and 7-day mortality were collected. RESULTS: A total of 37 cases were enrolled finally, including 17 cases with citric acid local anticoagulation and 20 cases with heparin systemic anticoagulation. There was no statistically significant difference in general data such as gender, age, and body weight of children between the two groups. There were no statistically significant differences in baseline levels of coagulation indicators and inflammatory mediators before treatment of children between the two groups. One day after weaning from CBP, both groups showed varying degrees of improvement in coagulation indicators compared with those before treatment. Compared with before treatment, the PT of the heparin systemic anticoagulation group was significantly shortened after 1 day of weaning (s: 11.82±2.05 vs. 13.64±2.54), APTT and TT were significantly prolonged [APTT (s): 51.54±12.69 vs. 35.53±10.79, TT (s): 21.95±4.74 vs. 19.30±3.33], D-dimer level was significantly reduced (mg/L: 1.92±1.58 vs. 4.94±3.94), with statistically significant differences (all P < 0.05). While in the citrate local anticoagulation group, only APTT was significantly prolonged after treatment compared with that before treatment (s: 49.28±10.32 vs. 34.34±10.32, P < 0.05). There were no statistically significant differences in other coagulation indicators compared with before treatment. Compared with the citric acid local anticoagulation group, the PT of the heparin systemic anticoagulation group was significantly shortened after treatment (s: 11.82±2.05 vs. 13.61±3.05, P < 0.05), and the D-dimer level was significantly reduced (mg/L: 1.92±1.58 vs. 3.77±2.38, P < 0.01). The levels of inflammatory mediators in both groups were significantly reduced 1 day after CBP weaning compared with those before treatment [citric acid local anticoagulation group: hs-CRP (mg/L) was 12.53±5.44 vs. 22.65±7.27, PCT (µg/L) was 1.86±1.20 vs. 3.30±2.34, IL-6 (ng/L) was 148.48±34.83 vs. 202.32±48.62, TNF-α (ng/L) was 21.38±7.71 vs. 55.14±15.07; heparin systemic anticoagulation group: hs-CRP (mg/L) was 11.82±4.93 vs. 21.62±8.35, PCT (µg/L) was 1.90±1.08 vs. 3.18±1.97, IL-6 (ng/L) was 143.81±33.41 vs. 194.02±46.89, TNF-α (ng/L) was 22.44±8.17 vs. 56.17±16.92, all P < 0.05]. However, there was no statistically significant difference between the two groups (all P > 0.05). There was no statistically significant difference in bleeding complication during CBP and 7-day mortality in children between the citrate local anticoagulation group and the heparin systemic anticoagulation group (5.9% vs. 30.0%, 17.6% vs. 20.0%, both P > 0.05). CONCLUSIONS: Heparin for systemic anticoagulation and regional citrate anticoagulation can significantly reduce the levels of IL-6, TNF-α, hs-CRP and PCT in children with septic shock, and relieve inflammatory storm. Compared with citric acid local anticoagulation, heparin systemic anticoagulation can shorten the PT and reduce the level of D-dimer in children with septic shock, which may benefit in the prevention and treatment of disseminated intravascular coagulation (DIC).

SFyNCS detects oncogenic fusions involving non-coding sequences in cancer.

Fusion genes are well-known cancer drivers. However, most known oncogenic fusions are protein-coding, and very few involve non-coding sequences due to lack of suitable detection tools. We develop SFyNCS to detect fusions of both protein-coding genes and non-coding sequences from transcriptomic sequencing data. The main advantage of this study is that we use somatic structural variations detected from genomic data to validate fusions detected from transcriptomic data. This allows us to comprehensively evaluate various fusion detection and filtering strategies and parameters. We show that SFyNCS has superior sensitivity and specificity over existing algorithms through extensive benchmarking in cancer cell lines and patient samples. We then apply SFyNCS to 9565 tumor samples across 33 tumor types in The Cancer Genome Atlas cohort and detect a total of 165,139 fusions. Among them, 72% of the fusions involve non-coding sequences. We find a long non-coding RNA to recurrently fuse with various oncogenes in 3% of prostate cancers. In addition, we discover fusions involving two non-coding RNAs in 32% of dedifferentiated liposarcomas and experimentally validated the oncogenic functions in mouse model.

SFyNCS detects oncogenic fusions involving non-coding sequences in cancer.

Fusion genes are well-known cancer drivers. However, very few known oncogenic fusions involve non-coding sequences. We develop SFyNCS with superior performance to detect fusions of both protein-coding genes and non-coding sequences from transcriptomic sequencing data. We validate fusions using somatic structural variations detected from the genomes. This allows us to comprehensively evaluate various fusion detection and filtering strategies and parameters. We detect 165,139 fusions in 9,565 tumor samples across 33 tumor types in the Cancer Genome Atlas cohort. Among them, 72% of the fusions involve non-coding sequences and many are recurrent. We discover two long non-coding RNAs recurrently fused with various partner genes in 32% of dedifferentiated liposarcomas and experimentally validated the oncogenic functions in mouse model.

Dose prediction of organs at risk in patients with cervical cancer receiving brachytherapy using needle insertion based on a neural network method.

OBJECTIVE: A neural network method was employed to establish a dose prediction model for organs at risk (OAR) in patients with cervical cancer receiving brachytherapy using needle insertion. METHODS: A total of 218 CT-based needle-insertion brachytherapy fraction plans for loco-regionally advanced cervical cancer treatment were analyzed in 59 patients. The sub-organ of OAR was automatically generated by self-written MATLAB, and the volume of the sub-organ was read. Correlations between D2cm3 of each OAR and volume of each sub-organ-as well as high-risk clinical target volume for bladder, rectum, and sigmoid colon-were analyzed. We then established a neural network predictive model of D2cm3 of OAR using the matrix laboratory neural net. Of these plans, 70% were selected as the training set, 15% as the validation set, and 15% as the test set. The regression R value and mean squared error were subsequently used to evaluate the predictive model. RESULTS: The D2cm3/D90 of each OAR was related to volume of each respective sub-organ. The R values for bladder, rectum, and sigmoid colon in the training set for the predictive model were 0.80513, 0.93421, and 0.95978, respectively. The ∆D2cm3/D90 for bladder, rectum, and sigmoid colon in all sets was 0.052 ± 0.044, 0.040 ± 0.032, and 0.041 ± 0.037, respectively. The MSE for bladder, rectum, and sigmoid colon in the training set for the predictive model was 4.779 × 10-3, 1.967 × 10-3 and 1.574 × 10-3, respectively. CONCLUSION: The neural network method based on a dose-prediction model of OAR in brachytherapy using needle insertion was simple and reliable. In addition, it only addressed volumes of sub-organs to predict the dose of OAR, which we believe is worthy of further promotion and application.

[MicroRNA-22-3p Regulates the Expression of Kruppel-like Factor 6 to Affect the Cardiomyocyte-like Differentiation of Bone Marrow Mesenchymal Stem Cell].

Objective To explore the effect of microRNA-22-3p (miR-22-3p) regulating the expression of Kruppel-like factor 6 (KLF6) on the cardiomyocyte-like differentiation of bone marrow mesenchymal stem cell (BMSC). Methods Rat BMSC was isolated and cultured,and the third-generation BMSC was divided into a control group,a 5-azacytidine(5-AZA)group,a mimics-NC group,a miR-22-3p mimics group,a miR-22-3p mimics+pcDNA group,and a miR-22-3p mimics+pcDNA-KLF6 group.Real-time fluorescent quantitative PCR (qRT-PCR) was carried out to determine the expression of miR-22-3p and KLF6 in cells.Immunofluorescence staining was employed to detect the expression of Desmin,cardiac troponin T (cTnT),and connexin 43 (Cx43).Western blotting was employed to determine the protein levels of cTnT,Cx43,Desmin,and KLF6,and flow cytometry to detect the apoptosis of BMSC.The targeting relationship between miR-22-3p and KLF6 was analyzed by dual luciferase reporter gene assay. Results Compared with the control group,5-AZA up-regulated the expression of miR-22-3p (q=7.971,P<0.001),Desmin (q=7.876,P<0.001),cTnT (q=10.272,P<0.001),and Cx43 (q=6.256,P<0.001),increased the apoptosis rate of BMSC (q=12.708,P<0.001),and down-regulated the mRNA (q=20.850,P<0.001) and protein (q=11.080,P<0.001) levels of KLF6.Compared with the 5-AZA group and the mimics-NC group,miR-22-3p mimics up-regulated the expression of miR-22-3p (q=3.591,P<0.001;q=11.650,P<0.001),Desmin (q=5.975,P<0.001;q=13.579,P<0.001),cTnT (q=7.133,P<0.001;q=17.548,P<0.001),and Cx43 (q=4.571,P=0.037;q=11.068,P<0.001),and down-regulated the mRNA (q=7.384,P<0.001;q=28.234,P<0.001) and protein (q=4.594,P=0.036;q=15.945,P<0.001) levels of KLF6.The apoptosis rate of miR-22-3p mimics group was lower than that of 5-AZA group (q=8.216,P<0.001).Compared with the miR-22-3p mimics+pcDNA group,miR-22-3p mimics+pcDNA-KLF6 up-regulated the mRNA(q=23.891,P<0.001) and protein(q=13.378,P<0.001)levels of KLF6,down-regulated the expression of Desmin (q=9.505,P<0.001),cTnT (q=10.985,P<0.001),and Cx43 (q=8.301,P<0.001),and increased the apoptosis rate (q=4.713,P=0.029).The dual luciferase reporter gene experiment demonstrated that KLF6 was a potential target gene of miR-22-3p. Conclusion MiR-22-3p promotes cardiomyocyte-like differentiation of BMSC by inhibiting the expression of KLF6.

FOXP1 inhibits pancreatic cancer growth by transcriptionally regulating IRF1 expression.

FOXP1, known as a Forkhead-box (FOX) family protein, plays an important role in human tumorigenesis. However, the function and molecular mechanism of FOXP1 in pancreatic cancer (PC) remain unclear. Here, we report that PC patients with FOXP1 overexpression had a higher survival rate compared to patients with low- FOXP1 expression. Additionally, high expression of FOXP1 can markedly inhibit the growth of pancreatic cancer in vivo and in vitro, whereas low expression of FOXP1 effectively promoted the tumorigenesis. Mechanistically, FOXP1 could directly bind the IRF1 promoter, which triggered the transcriptional activity of IRF1. Taken together, FOXP1 suppressed PC growth via IRF1-dependent manner, serving as a potential prognostic biomarker for patients with PC.

Potential lipid-based strategies of amphotericin B designed for oral administration in clinical application.

Amphotericin B (AmB) is regarded as a first-line therapy against life-threatening invasive fungal infections. Due to its poor oral bioavailability, AmB is restricted to intravenous administration in clinical practice. As science continues to move forward, two lipid-based formulations are successfully developed for oral AmB administration, currently undergoing phase I clinical trials. Encouragingly, lipid-AmB conjugates with emulsions also exhibit a better bioavailability, which may be another strategy to design oral AmB formulation in clinical practice. Thus, this review mainly focused on the two lipid-based formulations in clinical trials, and discussed the potential perspectives of AmB-lipid conjugation-loaded nanocochleates and emulsions.

Realization of thousand-second improved confinement plasma with Super I-mode in Tokamak EAST.

Mastering nuclear fusion, which is an abundant, safe, and environmentally competitive energy, is a great challenge for humanity. Tokamak represents one of the most promising paths toward controlled fusion. Obtaining a high-performance, steady-state, and long-pulse plasma regime remains a critical issue. Recently, a big breakthrough in steady-state operation was made on the Experimental Advanced Superconducting Tokamak (EAST). A steady-state plasma with a world-record pulse length of 1056 s was obtained, where the density and the divertor peak heat flux were well controlled, with no core impurity accumulation, and a new high-confinement and self-organizing regime (Super I-mode = I-mode + e-ITB) was discovered and demonstrated. These achievements contribute to the integration of fusion plasma technology and physics, which is essential to operate next-step devices.

TRAIL inhibition by soluble death receptor 5 protects against acute myocardial infarction in rats.

Acute myocardial infarction (AMI) is associated with high morbidity and mortality. An effective therapeutic strategy is to rescue cardiomyocytes from death. Apoptosis is a key reason of cardiomyocyte death that can be prevented. In this study, we investigated the role of TNF-related apoptosis-inducing ligand (TRAIL) in initiating apoptosis by binding to death receptor 5 (DR5), and this procession is inhibited by soluble DR5 (sDR5) in rats after AMI. First, we found that the level of TRAIL in serum was down-regulated in AMI patients. Then, TRAIL and DR5 expression was analysed in the myocardium of rats after AMI, and their expression was up-regulated. sDR5 treatment reduced the myocardial infarct size and the levels of CK-MB and cTn-I in serum. The expression of caspase 3 and PARP is decreased, but the anti-apoptotic factor Bcl-2 was increased in sDR5 treatment rats after AMI. DR5 expression was also analysed after sDR5 treatment and it was down-regulated, and a low level of DR5 expression seemed to be beneficial for the myocardium. Overall, our findings indicated that sDR5 decreases myocardial damage by inhibiting apoptosis in rat after AMI. We expect to observe the potential therapeutic effects of sDR5 on AMI in the future.

Clinical Significance of a Circulating Tumor Cell-based Classifier in Stage IB Lung Adenocarcinoma: A Multicenter, Cohort Study.

OBJECTIVE: To investigate the effectiveness of a CTC-based classifier in stratifying stage IB LUAD. SUMMARY OF BACKGROUND DATA: Stage IB LUADs have an approximately 70% 5-year survival rate. The clinical application of ACT is controversial due to inconsistent results in a series of trials and few useful guide biomarkers. Thus, there is a pressing need for robust biomarkers to stratify stage IB patients to define which group would most likely benefit from ACT. Methods: Two hundred twelve stage IB LUAD patients were enrolled and were divided into 3 independent cohorts. The aptamer-modified NanoVelcro system was used to enrich the CTCs. RESULTS: A cutoff of <4 or >4 CTCs as the optimal prognostic threshold for stage IB LUAD was generated to stratify the patients in a 70-patient cohort into low-risk and high-risk groups. Patients with ≥ 4 CTCs in the training cohort had shorter progression-free survival ( P < 0.0001) and overall survival ( P < 0.0001) than patients with <4 CTCs. CTC number remained the strongest predictor of progression-free survival and overall survival even in a multivariate analysis including other clinicopathological parameters. Furthermore, a nomogram based on the CTC count was developed to predict the 3-year and 5-year survival in the training cohort and performed well in the other 2 validation cohorts (C-index: 0.862, 0.853, and 0.877). CONCLUSION: The presence of >4 CTCs can define a high-risk subgroup, providing a new strategy to make optimal clinical decisions for stage IB LUAD.

Piceatannol protects rat neuron cells from oxygen-glucose deprivation reperfusion injury via regulation of GSK-3ß/Nrf2 signaling pathway.

OBJECTIVE: To investigate the effect and mechanism of piceatannol on cerebral ischemia-reperfusion injury. METHODS: The oxygen-glucose deprivation reperfusion (OGD/R) model was constructed in primary cultured suckling rat cortical neuron cells. After 2 h of oxygen-glucose deprivation, the cells were treated with piceatannol for 24 h. The cell survival rate was detected by MTT assay, and the degree of cell damage was detected by intracellular lactate dehydrogenase (LDH) release assay. The activity of superoxide dismutase (SOD) and the content of adenosine triphosphate (ATP) were detected by colorimetric method. The content of reactive oxygen species (ROS) was detected by flow cytometry or observed with inverted fluorescence microscope. The ultrastructure of mitochondria was observed with transmission electron microscopy. Western blotting was used to detect the phosphorylation levels of protein kinase B (AKT) and glycogen synthase kinase (GSK)-3ß. Immunofluorescence staining was used to observe the nuclear localization of nuclear factor-erythroid 2-related factor (Nrf) 2. After OGD/R neuron cells were pretreated with Nrf2 inhibitor ML385 for 24 h, the effect of Nrf2 on the improvement of cell activity and antioxidant activity of piceatannol were investigated. Western blotting was used to detect the protein expression levels of Nrf2, heme oxygenase (HO) 1 and NADPH quinone oxidoreductase (NQO) 1. RESULTS: Piceatannol significantly increased the survival rate of OGD/R neurons, decreased LDH release and reactive oxygen species content, increased SOD activity, ameliorated mitochondrial ultrastructural damage, increased mitochondrial membrane potential and ATP level (all P<0.05), increased phosphorylation of AKT and GSK-3ß protein, up-regulated the expression of Nrf2, HO-1 and NQO1 protein, increased the nuclear-to-plasma ratio of Nrf2, and promoted the nuclear transfer of Nrf2 (all P<0.05). ML385 could significantly reverse the rescue effect of paclitaxel on the model cells and the regulatory activities of SOD, ROS and LDH (all P<0.05). CONCLUSION: Piceatannol can regulate Nrf2 by activating GSK-3ß signaling pathway, promote its nuclear translocation, exert corresponding antioxidant effect, and protect mitochondrial structure and function in rat neuron cells.

Pinus massoniana needle extracts attenuate oxidative stress injury in cerebral ischemia reperfusion rats by regulating JNK3/caspase-3 signal transduction.

OBJECTIVE: To investigate the effect and mechanism of Pinus massoniana needle extracts (PNE) on oxidative stress injury in cerebral ischemia reperfusion rats. METHODS: The SD male rats were randomly divided into sham group, model control group, Edaravone (3 mg/kg) group, PNE low-dose (200 mg/kg), medium-dose (400 mg/kg) and high-dose (800 mg/kg) groups. PNE was administered by gavage for 7 d before modeling and 6 h after modeling in PNE treatment groups; Edaravone was given by intraperitoneal injection 7 d before modeling and 6 h after reperfusion. The rat model of cerebral ischemia reperfusion injury was established by middle cerebral artery occlusion method. After 24 h of reperfusion, the neurological deficit score, brain water content and cerebral infarction volume of rats were measured. The pathological changes of cerebral cortex and hippocampus were observed by HE staining, and the number of normal nerve cells was counted. The apoptosis rate of neurons in cerebral cortex was detected by TUNEL method. The content of nitric oxide (NO), malondialdehyde (MDA) and superoxide dismutase (SOD) activity in ischemic brain tissue were detected. The protein expression of c-Jun N-terminal kinase (JNK) 3, phosphorylated JNK3 (p-JNK3), B-cell lymphoma protein(Bcl) -2, Bcl-2 associated X (Bax), cytochrome C and caspase-3 in cerebral cortex were detected by Western blotting method. RESULTS: Compared with the model control group, the behavioral score, brain water content and cerebral infarction volume in PNE groups were significantly reduced (all P<0.05), the pathological damage of cerebral cortex and hippocampal CA1 area was significantly alleviated, and the number of normal nerve cells in ischemic cortex and hippocampal CA1 area was increased (all P<0.05). The medium-dose PNE group had the best effect. Compared with the model control group, the apoptosis rate of cortical neurons, the content of NO and MDA in cerebral cortex, the ratio of p-JNK3/JNK3, the expression level of cytochrome C and caspase-3 protein in PNE medium-dose group were significantly reduced , and the activity of SOD, the Bcl-2/Bax ratio were significantly improved (all P<0.05). CONCLUSION: PNE ameliorates brain injury after cerebral ischemia reperfusion in rats, which may be related to scavenging NO and MDA, inhibiting oxidative stress-mediated JNK3/caspase-3 signsal transduction to inhibit neuronal apoptosis.

Soluble Flt-1 in AMI Patients Serum Inhibits Angiogenesis of Endothelial Progenitor Cells by Suppressing Akt and Erk's Activity.

In acute myocardial infarction (AMI), endothelial progenitor cells (EPCs) are essential for the recovery of collateral circulation via angiogenesis. Clinical research has shown that the poor prognosis of the patients with AMI is closely associated with the cell quantity and function of EPCs. Whether there are differences in the biological features of EPCs from AMI patients and healthy subjects is worth exploring. In this study, EPCs were isolated from human peripheral blood and identified as late-stage EPCs by flow cytometry, immunofluorescence, and blood vessel formation assay. Compared to healthy subjects, AMI patients had more EPCs in the peripheral blood compared to healthy subjects. In addition, EPCs from AMI patients exhibited higher migration ability in the transwell assay compared to EPCs from healthy subjects. However, no difference in the angiogenesis of EPCs was observed between AMI patients and healthy subjects. Further studies revealed that soluble vascular endothelial growth factor receptor 1 (sFlt-1) in the serum of AMI patients was involved in the inhibition of EPCs angiogenesis by suppressing the Akt and Erk pathways. In conclusion, this study demonstrated that elevated serum sFlt-1 inhibits angiogenesis of EPC in AMI patients. Our findings uncover a pathogenic role of sFlt-1 in AMI.

Predictive Value of Circulating Tumor Cells Based on Subtraction Enrichment for Recurrence Risk in Stage II Colorectal Cancer.

Current guidelines recommend adjuvant chemotherapy (ACT) for stage II colorectal cancer (CRC) patients by using clinical high risk factors, with which circulating tumor cells (CTCs) were not considered. Here, an assessment to detect CTCs based on subtraction enrichment mediated by magnetic beads conjugated with CD45, immunofluorescence staining of CK, and fluorescence in situ hybridization of CEP8 is established. Both CEP8- and CK-positive CTCs have the potential to improve the risk stratification of stage II CRC patients. Patients with preoperative CTCs of ≥4 had a significantly higher recurrence risk than those with preoperative CTCs of <4 in two external validation cohorts (P < 0.0001). In the subgroup with clinical high risk, when preoperative CTCs were <4, patients did not benefit from ACT (P = 0.5764); however, when preoperative CTCs were ≥4, patients received benefit from ACT (P = 0.0064). Additionally, regardless of clinical risk status and preoperative CTC levels, if postoperative CTC levels were ≥4 for more than three consecutive time points (monitoring time interval, 2-6 months), the recurrence rate was 100%. Our findings suggested that the subtraction enrichment of CTCs could provide a reliable method to stratify the recurrence risk and make therapeutic decisions after surgery in stage II CRC patients.

Starfish infers signatures of complex genomic rearrangements across human cancers.

Complex genomic rearrangements (CGRs) are common in cancer and are known to form via two aberrant cellular structures-micronuclei and chromatin bridges. However, which of these mechanisms is more relevant to CGR formation in cancer and whether there are other undiscovered mechanisms remain unknown. Here we developed a computational algorithm, 'Starfish', to analyze 2,014 CGRs from 2,428 whole-genome-sequenced (WGS) tumors and discovered six CGR signatures based on their copy number and breakpoint patterns. Extensive benchmarking showed that our CGR signatures are highly accurate and biologically meaningful. Three signatures can be attributed to known biological processes-micronuclei- and chromatin-bridge-induced chromothripsis and circular extrachromosomal DNA. Over half of the CGRs belong to the remaining three signatures, not reported previously. A unique signature, which we named 'hourglass chromothripsis', with localized breakpoints and a low amount of DNA loss, is abundant in prostate cancer. Hourglass chromothripsis is associated with mutant SPOP, which may induce genome instability.

Evaluation of the effects of 1,25VitD3 on Th17 cells and Tregs in HTLV-1 infected cell lines.

Human T-lymphotropic virus type 1 (HTLV-1) causes chronic infections of human T lymphocytes. The present study aimed to evaluate the effects of 1,25VitD3 on the proportion of Tregs and Th17 cells, the expression of related transcription factors (ROR-γt and FOXP3) and cytokines (IL-10, TGF-ß, IL-6, and IL-17 A) in the HTLV-1infected cell lines MT-2 and MT-4. MT-2 and MT-4 cells and control PBMCs were treated with 1,25VitD3 and percentages of Tregs and Th17 cells was determined by flow cytometry. Gene expression and cytokine levels were analyzed by real-time PCR and ELISA, respectively. Treatment with-1,25VitD3 increased the percentage of Tregs in MT-2 and MT-4 cells, while it decreased the percentage of Th17 cells among MT-2 cells. 1,25VitD3 treatment also significantly improved FOXP3 gene expression in MT-2 cells, while reducing ROR-γt-gene expression in MT-2 and MT-4 cells comparing to untreated cells. Treatment with 1,25VitD3 significantly improved IL-10 levels in MT-2 cells, as well as TGF-ß levels in both cell lines culture supernatants. 1,25VitD3 treatment diminished IL-6 levels in cell culture supernatants of MT-2 and MT-4 as well as IL-17 A levels in MT-2. Here we showed, that 1,25VitD3 modulated immune responses by enhancing Tregs differentiation and functions as well as inhibiting Th17 differentiation and actions in HTLV-1 infected cell lines. This suggests that VitD3 may have therapeutic effects in HTLV-1-related diseases by suppressing adverse inflammatory responses. Keywords: Tregs; Th17 cells; HTLV-1; 1, 25VitD3.