Enhanced exon skipping and prolonged dystrophin restoration achieved by TfR1-targeted delivery of antisense oligonucleotide using FORCE conjugation in mdx mice.

Current therapies for Duchenne muscular dystrophy (DMD) use phosphorodiamidate morpholino oligomers (PMO) to induce exon skipping in the dystrophin pre-mRNA, enabling the translation of a shortened but functional dystrophin protein. This strategy has been hampered by insufficient delivery of PMO to cardiac and skeletal muscle. To overcome these limitations, we developed the FORCETM platform consisting of an antigen-binding fragment, which binds the transferrin receptor 1, conjugated to an oligonucleotide. We demonstrate that a single dose of the mouse-specific FORCE-M23D conjugate enhances muscle delivery of exon skipping PMO (M23D) in mdx mice, achieving dose-dependent and robust exon skipping and durable dystrophin restoration. FORCE-M23D-induced dystrophin expression reached peaks of 51%, 72%, 62%, 90% and 77%, of wild-type levels in quadriceps, tibialis anterior, gastrocnemius, diaphragm, and heart, respectively, with a single 30 mg/kg PMO-equivalent dose. The shortened dystrophin localized to the sarcolemma, indicating expression of a functional protein. Conversely, a single 30 mg/kg dose of unconjugated M23D displayed poor muscle delivery resulting in marginal levels of exon skipping and dystrophin expression. Importantly, FORCE-M23D treatment resulted in improved functional outcomes compared with administration of unconjugated M23D. Our results suggest that FORCE conjugates are a potentially effective approach for the treatment of DMD.

The biggest problem confronting oligonucleotide therapeutics is a lack of compounds capable of targeting compounds to diseased tissues. This paper reports a major advance targeting the transferrin receptor to increase the delivery of morpholine oligomers to muscle cells in vivo. This work suggests the possibility for improved treatments of muscular dystrophy and other diseases.

Homogeneous antibody and CAR-T cells with improved effector functions targeting SSEA-4 glycan on pancreatic cancer.

Pancreatic cancer is usually asymptomatic in the early stages; the 5-y survival rate is around 9%; and there is a lack of effective treatment. Here we show that SSEA-4 is more expressed in all pancreatic cancer cell lines examined but not detectable in normal pancreatic cells; and high expression of SSEA-4 or the key enzymes B3GALT5 + ST3GAL2 associated with SSEA-4 biosynthesis significantly lowers the overall survival rate. To evaluate potential new treatments for pancreatic cancer, homogeneous antibodies with a well-defined Fc glycan for optimal effector functions and CAR-T cells with scFv construct designed to target SSEA-4 were shown highly effective against pancreatic cancer in vitro and in vivo. This was further supported by the finding that a subpopulation of natural killer (NK) cells isolated by the homogeneous antibody exhibited enhancement in cancer-cell killing activity compared to the unseparated NK cells. These results indicate that targeting SSEA-4 by homologous antibodies or CAR-T strategies can effectively inhibit cancer growth, suggesting SSEA-4 as a potential immunotherapy target for treating pancreatic disease.

CDK4/6 inhibitors, PI3K/mTOR inhibitors, and HDAC inhibitors as second-line treatments for hormone receptor-positive, HER2-negative advanced breast cancer: a network meta-analysis.

BACKGROUND: This study sought to compare the benefits and safety of agents including Cyclin-dependent kinase 4/6 (CDK4/6) inhibitors, phosphoinositide 3-kinase (PI3K)/mammalian target of rapamycin (mTOR) inhibitors, and histone deacetylase (HDAC) inhibitors as second-line treatments for these patients by conducting a comprehensive systematic review and network meta-analysis. METHODS: The Medline, Embase and Cochrane Library databases were searched for randomized trials comparing CDK4/6 inhibitors, PI3K/mTOR inhibitors, or HDAC inhibitors vs. placebo with the addition of exemestane or fulvestrant as second-line treatments in patients with HR + advanced breast cancer up to December 16, 2021. Outcomes of interest were progression-free survival (PFS), overall response rate (ORR), overall survival (OS), clinical benefit rate (CBR), and grade 3-4 adverse drug events (ADEs). The present study was conducted according to the Cochrane Collaboration and PRISMA statements. The overall effect was pooled using the random effects model. RESULTS: Seventeen studies with a total of 9,100 participants were included in the current study. Compared with placebo plus fulvestrant, PFS was significantly improved by CDK4/6 inhibitor plus fulvestrant, mTOR inhibitor plus fulvestrant, mTOR inhibitor plus exemestane, and PI3K inhibitor plus fulvestrant, but not HDAC inhibitor plus exemestane. While mTOR inhibitor plus exemestane was the best regimen (SUCRA value 89.5%), the mTOR inhibitor plus exemestane regimen induced more severe adverse events (SAEs) than the HDAC inhibitor plus exemestane regimen [OR, 95% CI: 2.40 (1.40-4.10)]. CONCLUSION: mTOR inhibitor and CDK4/6 inhibitor-based regimens demonstrated superior clinical efficacy and comparable safety profiles as second-line treatment in patients with HR-positive, HER2-negative advanced breast cancer.

Analysis and identification of ferroptosis-related genes in ulcerative colitis.

BACKGROUND: Previous studies have shown that ferroptosis is associated with the pathogenesis of ulcerative colitis (UC). Therefore, this study aimed to identify key ferroptosis-related genes (FRGs) associated with the diagnosis of UC. METHODS: UC-related expression datasets were downloaded from the Gene Expression Omnibus (GEO) database. First, Weighted Gene Co-expression Network Analysis (WGCNA) was used to identify UC-related genes (UCRGs). Differentially expressed genes (DEGs) between normal and UC groups were screened in GSE87466, and DEGs were subjected to an intersection analysis with FRGs and UCRGs to obtain ferroptosis-related DEGs (FR DEGs). Then a protein-protein interaction (PPI) network was constructed for FR DEGs. The hub genes were extracted based on the degree, Maximum Neighborhood Component (MNC), closeness, and Maximal Clique Centrality (MCC). Biomarkers with diagnostic values were screened by support vector machine (SVM) and the least absolute shrinkage and selection operator (LASSO) algorithms. Next, the infiltration of immune cells was compared between UC and normal groups, and the correlation between different immune cells and diagnostic genes was analyzed. The biological functions, classical pathways, and intermolecular interaction networks of diagnostic genes were characterized utilizing ingenuity pathway analysis (IPA). Finally, a TF-mRNA network was constructed and potential small-molecule compounds were screened. RESULTS: Thirty-six FR DEGs were obtained, and these were enriched in biological processes such as positive regulation of cytokine production, cytokine-mediated signalling pathway, long-chain fatty acid-CoA ligase activity, etc. Among 18 hub genes, five genes (ALOX5, TIMP1, TNFAIP3, SOCS1, DUOX2) were captured with diagnostic values for UC, and they displayed significant differences between UC and normal groups. Sixteen immune cell infiltrates were significantly different between UC and normal groups, such as activated dendritic cells and resting dendritic cells. TNFAIP3 and ALOX5 were positively correlated with neutrophils, and TIMP1, SOCS1, ALOX5, and DUOX2 were negatively correlated with M2 macrophages. IPA showed that diagnostic genes were related to 43 function modules and activated 17 pathways. The constructed TF-mRNA regulatory network comprised three diagnostic genes and 17 differentially expressed TFs. Potential small-molecule compounds including helveticoside and cymarin were identified. CONCLUSION: Our findings yielded several promising FRGs for UC, providing a scientific reference for further studies on the pathogenesis of UC.

Development of 5-FU-modified tumor suppressor microRNAs as a platform for novel microRNA-based cancer therapeutics.

MicroRNA (miRNAs) are pleiotropic post-transcriptional modulators of gene expression. Their inherently pleiotropic nature makes miRNAs strong candidates for the development of cancer therapeutics, yet despite their potential, there remains a challenge to deliver nucleic acid-based therapies into cancer cells. We developed a novel approach to modify miRNAs by replacing the uracil bases with 5-fluorouracil (5-FU) in the guide strand of tumor suppressor miRNAs, thereby combining the therapeutic effect of 5-FU with tumor-suppressive effect of miRNAs to create a potent, multi-targeted therapeutic molecule without altering its native RNAi function. To demonstrate the general applicability of this approach to other tumor-suppressive miRNAs, we screened a panel of 12 novel miRNA mimetics in several cancer types, including leukemia, breast, gastric, lung, and pancreatic cancer. Our results show that 5-FU-modified miRNA mimetics have increased potency (low nanomolar range) in inhibiting cancer cell proliferation and that these mimetics can be delivered into cancer cells without delivery vehicle both in vitro and in vivo, thus representing significant advancements in the development of therapeutic miRNAs for cancer. This work demonstrates the potential of fluoropyrimidine modifications that can be broadly applicable and may serve as a platform technology for future miRNA and nucleic acid-based therapeutics.

Theoretical and numerical modeling of Rayleigh wave scattering by an elastic inclusion.

This work presents theoretical and numerical models for the backscattering of two-dimensional Rayleigh waves by an elastic inclusion, with the host material being isotropic and the inclusion having an arbitrary shape and crystallographic symmetry. The theoretical model is developed based on the reciprocity theorem using the far-field Green's function and the Born approximation, assuming a small acoustic impedance difference between the host and inclusion materials. The numerical finite element (FE) model is established to deliver a relatively accurate simulation of the scattering problem and to evaluate the approximations of the theoretical model. Quantitative agreement is observed between the theoretical model and the FE results for arbitrarily shaped surface/subsurface inclusions with isotropic/anisotropic properties. The agreement is excellent when the wavelength of the Rayleigh wave is larger than, or comparable to, the size of the inclusion, but it deteriorates as the wavelength gets smaller. Also, the agreement decreases with the anisotropy index for inclusions of anisotropic symmetry. The results lay the foundation for using Rayleigh waves for quantitative characterization of surface/subsurface inclusions, while also demonstrating its limitations.

Dual impact of ambient humidity on the virulence of Magnaporthe oryzae and basal resistance in rice.

Humidity is a critical environmental factor affecting the epidemic of plant diseases. However, it is still unclear how ambient humidity affects the occurrence of diseases in plants. In this study, we show that high ambient humidity enhanced blast development in rice plants under laboratory conditions. Furthermore, we found that high ambient humidity enhanced the virulence of Magnaporthe oryzae by promoting conidial germination and appressorium formation. In addition, the results of RNA-sequencing analysis and the ethylene content assessment revealed that high ambient humidity suppressed the accumulation of ethylene and the activation of ethylene signaling pathway induced by M. oryzae in rice. Knock out of ethylene signaling genes OsEIL1 and OsEIN2 or exogenous application of 1-methylcyclopropene (ethylene inhibitor) and ethephon (ethylene analogues) eliminated the difference of blast resistance between the 70% and 90% relative humidity conditions, suggesting that the activation of ethylene signaling contributes to humidity-modulated basal resistance against M. oryzae in rice. In conclusion, our results demonstrated that high ambient humidity enhances the virulence of M. oryzae and compromises basal resistance by reducing the activation of ethylene biosynthesis and signaling in rice. Results from this study provide cues for novel strategies to control rice blast under global environmental changes.

CEBPD modulates the airway smooth muscle transcriptomic response to glucocorticoids.

BACKGROUND: CCAAT/Enhancer Binding Protein D (CEBPD), a pleiotropic glucocorticoid-responsive transcription factor, modulates inflammatory responses. Of relevance to asthma, expression of CEBPD in airway smooth muscle (ASM) increases with glucocorticoid exposure. We sought to characterize CEBPD-mediated transcriptomic responses to glucocorticoid exposure in ASM by measuring changes observed after knockdown of CEBPD and its impact on asthma-related ASM function. METHODS: Primary ASM cells derived from four donors were transfected with CEBPD or non-targeting (NT) siRNA and exposed to vehicle control, budesonide (100 nM, 18 h), TNFα (10 ng/ml, 18 h), or both budesonide and TNFα. Subsequently, RNA-Seq was used to measure gene expression levels, and pairwise differential expression results were obtained for exposures versus vehicle and knockdown versus control conditions. Weighted gene co-expression analysis was performed to identify groups of genes with similar expression patterns across the various experimental conditions (i.e., CEBPD knockdown status, exposures). RESULTS: CEBPD knockdown altered expression of 3037 genes under at least one exposure (q-value < 0.05). Co-expression analysis identified sets of 197, 152 and 290 genes that were correlated with CEBPD knockdown status, TNFα exposure status, and both, respectively. JAK-STAT signaling pathway genes, including IL6R and SOCS3, were among those influenced by both TNFα and CEBPD knockdown. Immunoblot assays revealed that budesonide-induced IL-6R protein expression and augmented IL-6-induced STAT3 phosphorylation levels were attenuated by CEBPD knockdown in ASM. CONCLUSIONS: CEBPD modulates glucocorticoid responses in ASM, in part via modulation of IL-6 receptor signaling.

Transcription factor FOXC1 positively regulates SFRP1 expression in androgenetic alopecia.

Androgenetic alopecia (AGA) is the most common type of hair loss dysfunction. Secreted frizzled related protein 1 (SFRP1) is found to be associated with hair loss, but its role in AGA and the regulation mechanism of its transcription level is unclear. The aim of our study is to explore the expression of SFRP1 in AGA samples and its transcriptional mechanism. Male frontal and occipital scalp hair follicles from AGA patients were collected, and human dermal papilla cells (DPCs) were isolated and cultured. SFRP1 gene was cloned and constructed into recombinant plasmids to perform dual-luciferase reporter assay. Transcription factor binding sites were predicted through the Jaspar website and further confirmed by the chromatin immunoprecipitation (ChIP) assay. Expression of genes in DPCs was determined by immunofluorescence (IF) staining, quantitative real-time PCR (qRT-PCR) and western blotting. Our findings showed that SFRP1 was highly expressed in DPCs of AGA patients. The core promoter region of SFRP1 was from -100 to +50 bp and was found to be positively regulated by forkhead box C1 (FOXC1), a transcription factor related to hair growth, both at mRNA and protein level in DPCs. Our study suggests that FOXC1 plays an important role in regulating SFRP1 transcription, which may provide new insights into the development of therapeutic strategies for the treatment of AGA.

First Report of Leaf Spot on Stephania tetrandra Caused by Albifimbria verrucaria in China.

Stephania tetrandra S. Moore belongs to the family Menispermaceae and is a Chinese medicinal plant widely distributed in tropical and subtropical regions of Asia and Africa. The root can be used for a variety of treatments (Jiang et al. 2020). In August 2021, leaf spot symptoms were observed on S. tetrandra cultivated in Jiangxi (114.456E, 27.379N, southern China). The disease symptoms included a slight constriction of the leaves, with irregularly shaped brown to black spots with well-defined borders. Severely affected leaves were shed by the plant. In order to determine the cause, symptomatic leaves were surface-disinfested with 0.6% NaOCl for 2 min, and rinsed twice in sterile water, then incubated on moist paper towels at 26°C in the dark for 2 days. Cream-colored sporodochia were observed within the leaf spots, turning dark green to black within 16 hours. A slow-growing white fungus was isolated from 95% of the samples (n = 30) on PDA. Dark green sporodochia emerged after 7 to 10 days of incubation, and released tip-end oval, non-septate, hyaline conidia measuring 6.7 to 8.5 µm (mean 7.5 µm, n = 50) by 2.0 to 3.3 µm (mean 2.7 µm, n = 50). Concentric rings were interspersed with sporodochia on the continually incubated mycelium. The morphological characteristics of the isolates matched the description of Albifimbria (Lombard et al. 2016). Nucleotide sequences, amplified from isolate FJL5C using primers of the internal transcribed spacer (ITS) (White et al. 1990), calmodulin (cmdA; Carbone and Kohn 1999), and RNA polymerase II second largest subunit (rpb2; O'Donnell et al. 2007), were deposited in GenBank under accession numbers OM317911, OM386815, and OM386816. A BLASTn analysis of the sequences showed 100% identity with the type strain CBS 328.52 (Lombard et al. 2016) of Albifimbria verrucaria (syn. Myrothecium verrucaria) for ITS, and 99% for cmdA and rpb2 (KU845893, KU845875, and KU845931, respectively). A phylogenetic tree generated using the three sequences showed that the isolate from S. tetrandra grouped with the A. verrucaria isolates, but away from other species of Albifimbria. These results together with the lack of a pale luteus exudate produced by A. viridis (Lombard et al. 2016) implied that the isolate was A. verrucaria. The culture was deposited in Guangdong Microbial Culture Collection Center (GDMCC 3.716). To verify pathogenicity, conidial suspension (106 conidia/mL in 0.05% Tween 20 solution) was sprayed onto six healthy plants. Six other plants sprayed with the Tween 20 solution alone served as controls. All plants were incubated in the dark at 26°C and 95% humidity for 30 hours, then transferred to a greenhouse at 26°C and 12 hours of illumination per day for 2 to 3 days. Inoculated leaves developed similar symptoms to those described above, whereas control plants remained healthy. The same pathogen was isolated from the diseased leaves, with the same morphological and molecular traits as those from the field plants. This experiment fulfilled Koch's postulates and confirmed that A. verrucaria causes leaf spots on S. tetrandra. This pathogen has been reported to cause disease in a wide range of weeds, legumes, and crop plants (Herman et al. 2020). To our knowledge, this is the first report of A. verrucaria causing leaf spots on S. tetrandra in natural or controlled environments. The disease can seriously threaten S. tetrandra on growth and yield loss.

First Report of Sclerotium rolfsii Causing Southern Blight on Stephania tetrandra in China.

Stephania tetrandra S. Moore is a perennial liana and is widely cultivated in southern China for traditional Chinese medicine as a diuretic, anti-inflammatory, and antirheumatic treatment (Jiang et al. 2020). In August 2021, it was observed that a severe stem rot disease affected St. tetrandra cultivated in Anfu, Jiangxi province, China (114°27'26" E, 27°22'46" N). The disease symptoms included constriction and rot at the base of the stem, and covered with a layer of white mycelia. The plants above-ground finally wilted and dried with a disease incidence ranging from 8% to 16%. Lots of dried plants formed withered patches of field. Sections (1.0~2.0 cm) from browning stem tissues were surface-disinfected with 75% ethanol for 15 s, followed by 60 s in 4% NaClO, rinsed twice in sterile water, dried on sterilized filter paper, placed on potato dextrose agar (PDA), and incubated at 26°C in the dark for 3 days. A white rhizomorphic fungal mycelium, that is similar to the mycelium of strain FJSR0 on the surface of an infected plant in the field, was isolated from the cultured tissues with 67% frequency. When incubated on PDA, white and fluffy mycelia with even margins and a slight halo formed. Mycelia-produced clamp connections were observed. Colonies grew quickly and covered the dish (diameter: 9 cm) in 5 or 6 days. After that, sclerotia were initially white, then turned yellow, and chestnut brown at maturity. Spherical and subspherical sclerotia were observed after 8 days, with each plate containing 448 to 634 sclerotia (0.8 to 1.4 mm diameter; mean = 0.94 mm; n = 50). On the basis of morphology, the pathogen was similar to Sclerotium rolfsii Sacc. [teleomorph: Athelia rolfsii (Curzi) Tu & Kimbrough] (Sun et al. 2020; Ling et al. 2021). For molecular confirmation, the internal transcribed spacer (ITS) region with approximately 680 bp was amplified from strains FJRS0 and FJRS1 using primers ITS1/ITS4 (White et al. 1990). Two distinct types (different in one SNP and one 1-bp InDel) of ITS sequences were obtained from each isolate, and all isolates contain the two types (FJSR0: ON972516, ON972517; FJSR1: ON972520, ON972518). BLAST analysis of each type found that the hits, with identities >99%, are A. rolfsii except for two Sc. delphinii sequences (GU567775.1 and MK073010.1). Phylogenetic analysis placed strains FJSR0 and FJSR1 in the same clade as Sc. rolfsii but away from Sc. delphinii based on the previous method (Sun et al. 2021). Both morphological and molecular characteristics confirmed that the strains were Sc. rolfsii. For pathogenicity tests, PDA plugs (8 mm in diameter) covered with 5-day-old fungal mycelium were inoculated at the stem bases of three healthy St. tetrandra seedings and incubated at 26℃ and relative humidity of 80%. On the fifth day, inoculated plants were wilting. The infected stem bases turned brown to black and constricted as previously observed in the field. Some leaves, infected by the mycelium expanded from the PDA plugs, developed an orange and irregular spot. Sclerotia were observed 20 days post inoculation. In contrast, the leaves and stems of non-inoculated control plants remained symptomless. Pathogenicity tests were repeated three times. The fungus was reisolated consistently from each symptomatic tissue, thus completing Koch's postulates. Although Sc. rolfsii has been previously reported to cause a southern blight symptoms on vegetables, ornamentals, grass, and medicinal and leguminous crops (Sun et al. 2020; Ling et al. 2021), this is the first report of Sc. rolfsii causing similar symptoms of southern blight on St. tetrandra in China.

Clinical value of next-generation sequencing in guiding decisions regarding endocrine therapy for advanced HR-positive/HER-2-negative breast cancer.

Objective: The mechanism of acquired gene mutation plays a major role in resistance to endocrine therapy in hormone receptor (HR)-positive advanced breast cancer. Circulating tumor DNA (ctDNA) has been allowed for the assessment of the genomic profiles of patients with advanced cancer. We performed this study to search for molecular markers of endocrine therapy efficacy and to explore the clinical value of ctDNA to guide precise endocrine therapy for HR-positive/human epidermal growth factor receptor-2 (HER-2)-negative metastatic breast cancer patients. Methods: In this open-label, multicohort, prospective study, patients were assigned to four parallel cohorts and matched according to mutations identified in ctDNA: 1) activation of the phosphatidylinositol-3-kinase (PI3K)/AKT/mammalian target of rapamycin (mTOR) signaling pathway preferred mTOR inhibitor combined with endocrine therapy; 2) estrogen receptor 1 (ESR1) mutation preferred fulvestrant; 3) HER-2 mutations preferred pyrotinib; and 4) no actionable mutations received treatment according to the clinical situation. In all cohorts, patients were divided into compliance group and violation group. The primary outcome measure was progression-free survival (PFS), and the secondary outcome measure was overall survival (OS). Results: In all cohorts, the combined median PFS was 4.9 months, and median PFS for the compliance and violation groups was 6.0 and 3.0 months, respectively [P=0.022, hazard ratio (HR)=0.57]. Multivariate Cox regression model showed the risk of disease progression was lower in compliance group than in violation group (P=0.023, HR=0.55). Among the patients with HER-2 mutations, the median PFS was 11.1 months in the compliance group and 2.2 months in the violation group (P=0.011, HR=0.20). There was no significant difference in the median PFS between patients who did and did not comply with the treatment protocol in patients with activation of the PI3K/AKT/mTOR or ESR1 mutation. Conclusions: The results suggest that ctDNA may help to guide the optimal endocrine therapy strategy for metastatic breast cancer patients and to achieve a better PFS. Next-generation sequencing (NGS) detection could aid in distinguishing patients with HER-2 mutation and developing new treatment strategies.

Cataloguing experimentally confirmed 80.7 kb-long ACKR1 haplotypes from the 1000 Genomes Project database.

BACKGROUND: Clinically effective and safe genotyping relies on correct reference sequences, often represented by haplotypes. The 1000 Genomes Project recorded individual genotypes across 26 different populations and, using computerized genotype phasing, reported haplotype data. In contrast, we identified long reference sequences by analyzing the homozygous genomic regions in this online database, a concept that has rarely been reported since next generation sequencing data became available. STUDY DESIGN AND METHODS: Phased genotype data for a 80.6 kb region of chromosome 1 was downloaded for all 2,504 unrelated individuals of the 1000 Genome Project Phase 3 cohort. The data was centered on the ACKR1 gene and bordered by the CADM3 and FCER1A genes. Individuals with heterozygosity at a single site or with complete homozygosity allowed unambiguous assignment of an ACKR1 haplotype. A computer algorithm was developed for extracting these haplotypes from the 1000 Genome Project in an automated fashion. A manual analysis validated the data extracted by the algorithm. RESULTS: We confirmed 902 ACKR1 haplotypes of varying lengths, the longest at 80,584 nucleotides and shortest at 1,901 nucleotides. The combined length of haplotype sequences comprised 19,895,388 nucleotides with a median of 16,014 nucleotides. Based on our approach, all haplotypes can be considered experimentally confirmed and not affected by the known errors of computerized genotype phasing. CONCLUSIONS: Tracts of homozygosity can provide definitive reference sequences for any gene. They are particularly useful when observed in unrelated individuals of large scale sequence databases. As a proof of principle, we explored the 1000 Genomes Project database for ACKR1 gene data and mined long haplotypes. These haplotypes are useful for high throughput analysis with next generation sequencing. Our approach is scalable, using automated bioinformatics tools, and can be applied to any gene.

Exosome-mediated pyroptosis of miR-93-TXNIP-NLRP3 leads to functional difference between M1 and M2 macrophages in sepsis-induced acute kidney injury.

Sepsis is a systemic inflammatory response syndrome caused by infection, resulting in organ dysfunction. Sepsis-induced acute kidney injury (AKI) is one of the most common potential complications. Increasing reports have shown that M1 and M2 macrophages both take part in the progress of AKI by influencing the level of inflammatory factors and the cell death, including pyroptosis. However, whether M1 and M2 macrophages regulate AKI by secreting exosome remains unknown. In the present study, we isolated the exosomes from M1 and M2 macrophages and used Western blot and enzyme-linked immunosorbent assay (ELISA) to investigate the effect of M1 and M2 exosomes on cell pyroptosis. miRNA sequencing was used to identify the different miRNA in M1 and M2 exosomes. Luciferase reporter assay was used to verify the target gene of miRNA. We confirmed that exosomes excreted by macrophages regulated cell pyroptosis in vitro by using Western blot and ELISA. miRNA sequencing revealed the differentially expressed level of miRNAs in M1 and M2 exosomes, among which miR-93-5p was involved in the regulation of pyroptosis. By using bioinformatics predictions and luciferase reporter assay, we found that thioredoxin-interacting protein (TXNIP) was a direct target of miR-93-5p. Further in vitro and in vivo experiments indicated that exosomal miR-93-5p regulated the TXNIP directly to influence the pyroptosis in renal epithelial cells, which explained the functional difference between different phenotypes of macrophages. This study might provide new targets for the treatment of sepsis-induced AKI.

Water resources allocation model based on ecological priority in the arid region.

Based on the characteristics of arid regions in the central and southern Ningxia，firstly we constructed the theoretical framework about rational allocation of water resources based on ecological priority, which supplemented and improved the theoretical system on rational allocation of water resources for sustainable development. On the basis of theoretical research, depending on its characteristic of water resources：pumping water from Yellow River in the north, piloting water from Jing River in the south and regional water resources, which formed a mixing water supply pattern of "pumping, piloting, storage". The theory of water resource allocation system was perfected. Then the mathematical model for the rational allocation of water resources was established on the basis of biggest benefit combined with economy, society and ecology. The model coincided the ecological water and domestic water benefit into the overall benefits, and provides them larger benefit coefficient and the priority order coefficient to realize the reasonable allocation of water resources on ecological priority mode. Finally, the results of allocation are analysed in detail, which provides constructive suggestions and guiding direction for the study of the rational allocation of regional water resources.

Circ_0091581 Promotes the Progression of Hepatocellular Carcinoma Through Targeting miR-591/FOSL2 Axis.

BACKGROUND: Circular RNAs (circRNAs) have shown crucial regulatory roles in cancer biology. We aimed to uncover the role and underlying mechanism of circ_0091581 in hepatocellular carcinoma (HCC) progression. METHODS: The abundance of circ_0091581, microRNA-591 (miR-591) and FOS like 2, AP-1 transcription factor subunit (FOSL2) was measured by quantitative real-time polymerase chain reaction. Cell viability, colony formation ability, and invasion ability were assessed by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, colony formation assay, and transwell invasion assay. The migration ability was analyzed by transwell migration assay and wound healing assay. Flow cytometry was used to evaluate the cell cycle and apoptosis of HCC cells. The interaction between miR-591 and circ_0091581 or FOSL2 was predicted by Circular RNA Interactome database or TargetScan database and confirmed by dual-luciferase reporter assay and RNA immune co-precipitation assay. FOSL2 protein expression was measured by Western blot assay. Xenograft tumor assay was conducted to analyze the role of circ_0091581 in HCC tumor growth in vivo. RESULTS: Circ_0091581 was highly expressed in HCC tissue samples and cell lines in contrast to that in adjacent normal tissue samples and THLE-2 cell line. Circ_0091581 accelerated the viability, colony formation, metastasis, and cell cycle, while it impeded the apoptosis of HCC cells. MiR-591 bound to circ_0091581, and circ_0091581 knockdown-mediated effects in HCC cells were largely overturned by miR-591 silencing. FOSL2 was a target of miR-591, and FOSL2 overexpression largely reversed miR-591 accumulation-induced influences in HCC cells. FOSL2 protein expression was down-regulated by circ_0091581 silencing, and the addition of miR-591 inhibitor partly recovered the expression of FOSL2 in HCC cells. Circ_0091581 interference notably suppressed HCC tumor growth in vivo. CONCLUSION: Circ_0091581 acted as an oncogene to enhance the viability, colony formation, metastasis and cell cycle and inhibit the apoptosis of HCC cells through targeting miR-591/FOSL2 axis.

In situ characterization of laser-generated melt pools using synchronized ultrasound and high-speed X-ray imaging.

Metal additive manufacturing is a fabrication method that forms a part by fusing layers of powder to one another. An energy source, such as a laser, is commonly used to heat the metal powder sufficiently to cause a molten pool to form, which is known as the melt pool. The melt pool can exist in the conduction or the keyhole mode where the material begins to rapidly evaporate. The interaction between the laser and the material is physically complex and difficult to predict or measure. In this article, high-speed X-ray imaging was combined with immersion ultrasound to obtain synchronized measurements of stationary laser-generated melt pools. Furthermore, two-dimensional and three-dimensional finite-element simulations were conducted to help explain the ultrasonic response in the experiments. In particular, the time-of-flight and amplitude in pulse-echo configuration were observed to have a linear relationship to the depth of the melt pool. These results are promising for the use of ultrasound to characterize the melt pool behavior and for finite-element simulations to aid in interpretation.

Clinical features and prognostic factors for extracranial oligometastatic breast cancer in China.

Evidence of an oligometastatic state in metastatic breast cancer (MBC) is relatively limited. The aim of our study was to investigate the clinical features and prognostic factors for extracranial oligometastatic breast cancer and to identify the best treatment approaches in this select population. Fifty postoperative inpatients diagnosed with extracranial oligometastatic breast cancer at the National Cancer Center in China between 2009 and 2014 were consecutively enrolled. Oligometastatic breast cancer was defined as MBC with three or fewer metastatic lesions confined to one organ; de novo Stage IV disease and local-regional recurrence were excluded. The median progression-free survival (PFS) and overall survival (OS) times were 15.2 and 78.9 months, respectively, and the 2-year PFS and 5-year OS rates were 40% and 58%, respectively. First-line treatment approach with standard systemic treatment + surgical resection for all metastatic lesions was an independent prognostic factor for prolonged PFS (hazard ratio = 0.32; 95% confidence interval [CI], 0.14-0.73; P = .006) and OS (hazard ratio = 0.35; 95% CI, 0.14-0.86; P = .022). Subgroup analysis showed that patients with a disease-free interval (DFI) ≥24 months, one metastatic lesion or the hormone receptor (HR) + subtype were more likely to get benefit from resection. Patients with oligometastatic breast cancer have a relatively good prognosis. Surgical resection for metastatic lesions could significantly improve PFS and OS. Further prospective research is warranted to confirm the results and to develop biomarkers for better patient selection.

Platinum-based chemotherapy in advanced triple-negative breast cancer: A multicenter real-world study in China.

Platinum-based chemotherapy (PBC) has proven benefits in phase III studies for advanced triple-negative breast cancer (TNBC) patients; however, real-world data of large samples from multiple centers are lacking. Our study was to compare the effectiveness of PBC and non-PBC in advanced TNBC patients in multicenter real-world settings. Totally, 495 patients with advanced TNBC receiving PBC (n = 350) or non-PBC (n = 145) at four cancer centers in China between 2003 and 2019 were included. Treatment responses and outcomes were compared between the two groups from first-line to third-line treatment. Of patients with PBC, 249 (71.1%) received PBC from first-line chemotherapy, 86 (24.6%) from second-line and 15 (4.3%) from third-line treatment. In first-line treatment, PBC was superior to non-PBC in objective response rate (ORR, 53.0% vs 32.1%, P < .001) and median progression-free survival (PFS, 8.4 vs 6.0 months, P = .022), whereas overall survival (OS) was similar (19.2 vs 16.8 months, P = .439). When comparing patients receiving non-PBC doublets (n = 221) with those receiving PBC doublets (n = 249), the same trend was observed in ORR (32.6% vs 53.0%, P < .001), median first-line PFS (6.5 vs 8.4 months, P = .041) and median first-line OS(17.8 vs 19.2 months, P = .568). Paclitaxel/docetaxel + platinum was more likely to be used, followed by gemcitabine + platinum. In second/third-line treatment, PBC yielded a similar response and survival compared to non-PBC. Adding PBC in the first-line therapy was better than that in the latter-line of treatment in terms of ORR, PFS and OS (P < .001). Toxic effects of PBC were tolerable and the most common adverse event was neutropenia (38.6%). PBC doublets exhibited superior efficacy and manageable toxicity compared to non-PBC doublets in the first-line treatment for Chinese mTNBC patients.

Unjamming and collective migration in MCF10A breast cancer cell lines.

Each cell comprising an intact, healthy, confluent epithelial layer ordinarily remains sedentary, firmly adherent to and caged by its neighbors, and thus defines an elemental constituent of a solid-like cellular collective [1,2]. After malignant transformation, however, the cellular collective can become fluid-like and migratory, as evidenced by collective motions that arise in characteristic swirls, strands, ducts, sheets, or clusters [3,4]. To transition from a solid-like to a fluid-like phase and thereafter to migrate collectively, it has been recently argued that cells comprising the disordered but confluent epithelial collective can undergo changes of cell shape so as to overcome geometric constraints attributable to the newly discovered phenomenon of cell jamming and the associated unjamming transition (UJT) [1,2,5-9]. Relevance of the jamming concept to carcinoma cells lines of graded degrees of invasive potential has never been investigated, however. Using classical in vitro cultures of six breast cancer model systems, here we investigate structural and dynamical signatures of cell jamming, and the relationship between them [1,2,10,11]. In order of roughly increasing invasive potential as previously reported, model systems examined included MCF10A, MCF10A.Vector; MCF10A.14-3-3ζ; MCF10.ErbB2, MCF10AT; and MCF10CA1a [12-15]. Migratory speed depended on the particular cell line. Unsurprisingly, for example, the MCF10CA1a cell line exhibited much faster migratory speed relative to the others. But unexpectedly, across different cell lines higher speeds were associated with enhanced size of cooperative cell packs in a manner reminiscent of a peloton [9]. Nevertheless, within each of the cell lines evaluated, cell shape and shape variability from cell-to-cell conformed with predicted structural signatures of cell layer unjamming [1]. Moreover, both structure and migratory dynamics were compatible with previous theoretical descriptions of the cell jamming mechanism [2,10,11,16,17]. As such, these findings demonstrate the richness of the cell jamming mechanism, which is now seen to apply across these cancer cell lines but remains poorly understood.