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Background & Aims: The expression of HBsAg from integrated HBV DNA limits the achievement of functional cure for chronic hepatitis B. Thus, characterising the unique expression and secretion of HBsAg derived from integrated HBV DNA is of clinical significance. Methods: A total of 563 treatment-naive patients and 62 functionally cured patients were enrolled, and HBsAg and HBcAg immunohistochemistry of their liver biopsy tissues was conducted followed by semi-quantitative analysis. Then, based on stratified analysis of HBeAg-positive and -negative patients, long-read RNA sequencing analysis, as well as an in vitro HBV integration model, we explored the HBsAg secretion characteristics of integrated HBV DNA and underlying mechanisms. Results: In contrast to the significantly lower serum HBsAg levels, no significant decrease of intrahepatic HBsAg protein was observed in HBeAg-negative patients, as compared with HBeAg-positive patients. The results of long-read RNA sequencing of liver tissues from patients with chronic HBV infection and in vitro studies using integrated HBV DNA mimicking dslDNA plasmid revealed that, the lower HBsAg secretion efficiency seen in HBeAg-negative patients might be attributed to an increased proportion of preS1 mRNA derived from integrated HBV DNA instead of covalently closed circular DNA. The latter resulted in an increased L-HBsAg proportion and impaired HBsAg secretion. Enhancer 1 (EnhI) in integrated HBV DNA could retarget preS1 (SP1) and preS2 (SP2) promoters to disrupt their transcriptional activity balance. Conclusions: The secretion of HBsAg originating from integrated HBV DNA was impaired. Mechanistically, functional deficiency of core promoter leads to retargeting of EnhI and thus uneven activation of the SP1 over the SP2 promoter, resulting in an increase in the proportion of L-HBsAg. Impact and implications: Integrated hepatitis B virus (HBV) DNA can serve as an important reservoir for HBV surface antigen (HBsAg) expression, and this limits the achievement of a functional cure. This study revealed that secretion efficiency is lower for HBsAg derived from integrated HBV DNA than HBsAg derived from covalently closed circular DNA, as determined by the unique sequence features of integrated HBV DNA. This study can broaden our understanding of the role of HBV integration and shed new light on antiviral strategies to facilitate a functional cure. We believe our results are of great general interest to a broad audience, including patients and patient organisations, the medical community, academia, the life science industry and the public.

Abstract CRISPR-based gene therapy offers precise targeting and specific editing of disease-related gene sequences, potentially yielding long-lasting treatment effects. However, efficient delivery remains a significant challenge for its widespread application. In this study, we design a novel short peptide-conjugated bioreducible polymer named TSPscp as a safe and effective delivery vector for the CRISPR system. Our results show that TSPscp markedly boosts transcriptional activation and genome editing activities of multiple CRISPR systems as confirmed by decomposition-seq and Deep-seq, which is resulted from its capability in facilitating delivery of plasmid DNA by promoting cellular uptake and lysosomal escape. Additionally, TSPscp further enhances genome editing of CRISPR by delivery of minicircle DNA, a condensed form of regular plasmid DNA. More importantly, TSPscp significantly improves delivery and genome editing of CRISPR system in vivo. In summary, our study highlights TSPscp as a promising delivery tool for CRISPR applications in vivo. Graphical Abstract

SRY‐box transcription factor 6 (SOX6) is a member of the SOX gene family and inhibits the proliferation of cervical cancer cells by inducing cell cycle arrest. However, the final cell fate and significance of these cell‐cycle‐arrested cervical cancer cells induced by SOX6 remains unclear. Here, we report that SOX6 inhibits the proliferation of cervical cancer cells by inducing cellular senescence, which is mainly mediated by promoting transforming growth factor beta 2 (TGFB2) gene expression and subsequently activating the TGFβ2–Smad2/3–p53–p21WAF1/CIP1–Rb pathway. SOX6 promotes TGFB2 gene expression through the MAP4K4–MAPK (JNK/ERK/p38)–ATF2 and WT1–ATF2 pathways, which is dependent on its high‐mobility group (HMG) domain. In addition, the SOX6‐induced senescent cervical cancer cells are resistant to cisplatin treatment. ABT‐263 (navitoclax) and ABT‐199 (venetoclax), two classic senolytics, can specifically eliminate the SOX6‐induced senescent cervical cancer cells, and thus significantly improve the chemosensitivity of cisplatin‐resistant cervical cancer cells. This study uncovers that the MAP4K4/WT1–ATF2–TGFβ2 axis mediates SOX6‐induced cellular senescence, which is a promising therapeutic target in improving the chemosensitivity of cervical cancer.

Abstract Background Breast cancer has the highest incidence rate of cancer worldwide, and brain metastases (BrM) are among the most malignant cases. While some patients have benefited from immune checkpoint inhibitors (ICIs), the complex anatomical structure of the brain and the heterogeneity of metastatic tumors have made it difficult to characterize the tumor immune microenvironment (TME) of metastatic tumors. Methods To address this, we used single‐cell RNA sequencing (scRNA‐seq) to analyze immune cells in the cerebrospinal fluid (CSF) of BrM patients with breast cancer, thereby providing a comprehensive view of the immune microenvironment landscape of BrM. Results Based on canonical marker genes, we identified nine cell types, and further identified their subtypes through differential expression gene (DEG) analysis. We compared the changes in cells and functions in the immune microenvironment of patients with different prognoses. Our analysis revealed a series of genes that promote tumor immune function (CCR5, LYZ, IGKC, MS4A1, etc.) and inhibit tumor immune function (SCGB2A2, CD24, etc.). Conclusions The scRNA‐seq in CSF provides a noninvasive method to describe the TME of breast cancer patients and guide immunotherapy.

ABSTRACTEbola virus (EBOV) belongs to Filoviridae family possessing single-stranded negative-sense RNA genome, which is a serious threat to human health. Nowadays, no therapeutics have been proven to be successful in efficiently decreasing the mortality rate. RNA binding proteins (RBPs) are reported to participate in maintaining cell integrity and regulation of viral replication. However, little is known about whether and how RBPs participate in regulating the life cycle of EBOV. In our study, we found that RNA binding motif protein 4 (RBM4) inhibited the replication of EBOV in HEK293T and Huh-7 cells by suppressing viral mRNA production. Such inhibition resulted from the direct interaction between the RRM1 domain of RBM4 and the “CU” enrichment elements located in the PE1 and TSS of the 3′-leader region within the viral genome. Simultaneously, RBM4 could upregulate the expression of some cytokines involved in the host innate immune responses to synergistically exert its antiviral function. The findings therefore suggest that RBM4 might serve as a novel target of anti-EBOV strategy.

Nipah virus (NiV), a highly pathogenic Henipavirus in humans, has been responsible for annual outbreaks in recent years. Experiments involving live NiV are highly restricted to biosafety level 4 (BSL-4) laboratories, which impedes NiV research. In this study, we developed transcription and replication-competent NiV-like particles (trVLP-NiV) lacking N, P, and L genes. This trVLP-NiV exhibited the ability to infect and continuously passage in cells ectopically expressing N, P, and L proteins while maintaining stable genetic characteristics. Moreover, the trVLP-NiV displayed a favourable safety profile in hamsters. Using the system, we found the NiV nucleoprotein residues interacting with viral RNA backbone affected viral replication in opposite patterns. This engineered system was sensitive to well-established antiviral drugs, innate host antiviral factors, and neutralizing antibodies. We then established a high-throughput screening platform utilizing the trVLP-NiV, leading to the identification of tunicamycin as a potential anti-NiV compound. Evidence showed that tunicamycin inhibited NiV replication by decreasing the infectivity of progeny virions. In conclusion, this trVLP-NiV system provided a convenient and versatile molecular tool for investigating NiV molecular biology and conducting antiviral drug screening under BSL-2 conditions. Its application will contribute to the development of medical countermeasures against NiV infections.

ABSTRACTThe persistence of HBV covalently closed circular DNA (cccDNA) and HBV integration into the host genome in infected hepatocytes pose significant challenges to the cure of chronic HBV infection. Although CRISPR/Cas9-mediated genome editing shows promise for targeted clearance of viral genomes, a safe and efficient delivery method is currently lacking. Here, we developed a novel approach by combining light-induced heterodimerization and protein acylation to enhance the loading efficiency of Cas9 protein into extracellular vesicles (EVs). Moreover, vesicular stomatitis virus-glycoprotein (VSV-G) was incorporated onto the EVs membrane, significantly facilitating the endosomal escape of Cas9 protein and increasing its gene editing activity in recipient cells. Our results demonstrated that engineered EVs containing Cas9/gRNA and VSV-G can effectively reduce viral antigens and cccDNA levels in the HBV-replicating and infected cell models. Notably, we also confirmed the antiviral activity and high safety of the engineered EVs in the HBV-replicating mouse model generated by hydrodynamic injection and the HBV transgenic mouse model. In conclusion, engineered EVs could successfully mediate functional CRISPR/Cas9 delivery both in vitro and in vivo, leading to the clearance of episomal cccDNA and integrated viral DNA fragments, and providing a novel therapeutic approach for curing chronic HBV infection.

ABSTRACT Monkeypox virus (MPXV) poses a global health threat. Droplet digital PCR (ddPCR) holds potential as an accurate diagnostic tool for clinical microbiology. However, there is limited literature on the applicability of ddPCR in clinical settings. In this study, the clinical features of patients with MPXV during the initial outbreak in China in June 2023 were reviewed, and an optimized ddPCR method with dilution and/or inhibitor removal was developed to enhance MPXV detection efficiency. Eighty-two MPXV samples were tested from nine different clinical specimen types, including feces, urine, pharyngeal swabs, anal swabs, saliva, herpes fluid, crust, and semen, and the viral load of each specimen was quantified. A comparative analysis was performed with qPCR to assess sensitivity and specificity and to investigate the characteristics of MPXV infection by analyzing viral loads in different clinical specimens. Consequently, common pharyngeal and gastrointestinal symptoms were observed in patients with MPXV. The optimized ddPCR method demonstrated relatively high sensitivity for MPXV quantification in the clinical materials, with a limit of detection of 0.1 copies/μL. This was particularly evident in low-concentration samples like whole blood, semen, and urine. The optimized ddPCR demonstrated greater detection accuracy compared with normal ddPCR and qPCR, with an area under the curve (AUC) of 0.939. Except for crust samples, viral loads in the specimens gradually decreased as the disease progressed. Virus levels in feces and anal swabs kept a high detection rate at each stage of post-symptom onset, and feces and anal swabs samples may be suitable for clinical diagnosis and continuous monitoring of MPXV.IMPORTANCEThe ddPCR technique proved to be a sensitive and valuable tool for accurately quantifying MPXV viral loads in various clinical specimen types. The findings provided valuable insights into the necessary pre-treatment protocols for MPXV diagnosis in ddPCR detection and the potentially suitable sample types for collection. Therefore, such results can aid in comprehending the potential characteristics of MPXV infection and the usage of ddPCR in clinical settings.

Abstract Background Government purchase of social forces to participate in old age care services can release the burden of social care. Current research on performance evaluation in this field mainly focussed on the establishment of appropriate evaluation indices. However, discussion on the policy implementation deviation is scarce. This study aimed to evaluate the performance of China’s local government purchase of old age care services, analyse the characteristics of related policies and explore their deviation. Methods The persons who participated in the Training of the Trainer (ToT) organized by the Red Cross Society were enrolled. The policy documents were obtained from the official websites. The K-means cluster was used to determine the project performance grades. We compared the project performance grades between service objects and undertakers with different characteristics utilizing the non-parametric test. Based on the framework of ‘Collaborative Participation – Project Performance Objective’, we analysed the content of relevant policy aiding by NVivo 12. Results Data of project performance were collected from 306 participants. The standardized mean score of the efficiency dimension was the lowest (0.70 ± 0.24). The projects were divided into four grades: poor (17.0%), average (27.5%), good (12.4%) and excellent (43.1%). There were statistically significant differences in project performance grades only between advanced ageing groups (Z = 2.429, P = 0.015). As well, the policy also mentioned that the services focus should be tilted towards the oldest old. The purchasers mainly involved the Ministry of Civil Affairs and Health management departments in the policy. Respite services were less mentioned in the responsibilities of the undertakers. The requirement for efficiency and effectiveness was mentioned in less than half of the policy documents. Conclusion Policy attention is needed for the responsibilities and functions of the intermediate purchasing force, as well as more precise directions and responsibilities of undertakers. The purchasers and undertakers should improve management abilities and capacity of old age care services and focus on associated factors to achieve the best marginal benefit. In addition, the embedded performance evaluation needs to be updated periodically to bridge the deviation between policy implementation and policy formulation.

Estimating the spatiotemporal variations in natural grassland carrying capacity is crucial for maintaining the balance between grasslands and livestock. However, accurately assessing this capacity presents significant challenges due to the high costs of biomass measurement and the impact of human activities. In this study, we propose a novel method to estimate grassland carrying capacity based on potential net primary productivity (NPP), applied to the source area of the Nujiang River and Selinco Lake on the Tibetan Plateau. Initially, we utilize multisource remote sensing data—including soil, topography, and climate information—and employ the random forest regression algorithm to model potential NPP in areas where grazing is banned. The construction of the random forest model involves rigorous feature selection and hyperparameter optimization, enhancing the model’s accuracy. Next, we apply this trained model to areas with grazing, ensuring a more accurate estimation of grassland carrying capacity. Finally, we analyze the spatiotemporal variations in grassland carrying capacity. The main results showed that the model achieved a high level of precision, with a root mean square error (RMSE) of 4.89, indicating reliable predictions of grassland carrying capacity. From 2001 to 2020, the average carrying capacity was estimated at 9.44 SU/km2, demonstrating a spatial distribution that decreases from southeast to northwest. A slight overall increase in carrying capacity was observed, with 65.7% of the area exhibiting an increasing trend, suggesting that climate change has a modest positive effect on the recovery of grassland carrying capacity. Most of the grassland carrying capacity is found in areas below 5000 m in altitude, with alpine meadows and alpine meadow steppes below 4750 m being particularly suitable for grazing. Given that the overall grassland carrying capacity remains low, it is crucial to strictly control local grazing intensity to mitigate the adverse impacts of human activities. This study provides a solid scientific foundation for developing targeted grassland management and protection policies.

A thorough understanding of the development process of grazing activities and an elucidation of their complex mechanisms are crucial for the formulation and adjustment of livestock management policies. In the Selinco region of the Qinghai–Tibet Plateau, we conducted a comprehensive analysis of year-end livestock numbers and densities over the past 30 years. The results indicate a gradual decline in overall livestock numbers in the Selinco region during this period, with a notable decrease between 2004 and 2014, followed by stabilization. Notably, the number of yaks has significantly increased, whereas the numbers of sheep, goats, and horses have markedly decreased. Regarding livestock density, there is a spatial pattern of decrease from northwest to southeast, with the density order being Seni District > Bange County > Anduo County > Shenzha County > Nima County > Shuanghu County. Between 2004 and 2014, most counties experienced a significant decline in livestock density, exhibiting a trough–peak pattern. However, after 2014, a complex spatiotemporal dynamic emerged. Concerning driving factors, from 1990 to 2004, rural population and economic development were the primary influences on livestock density. After 2004, forage–livestock balance management policies, snowstorms, and fluctuations in livestock prices likely became the main influencing factors. Further detailed analysis of these factors is essential for developing more effective management strategies.

Abstract Abnormal 5-methylcytosine (m5C) methylation has been proved to be closely related to gastric carcinogenesis, progression, and prognosis. Dysregulated long noncoding RNAs (lncRNAs) participate in a variety of biological processes in cancer. However, to date, m5C-methylated lncRNAs are rarely researched in gastric cancer (GC). Here, we found that RNA cytosine-C(5)-methyltransferase (NSUN2) was upregulated in GC and high NSUN2 expression was associated with poor prognosis. NR_033928 was identified as an NSUN2-methylated and upregulated lncRNA in GC. Functionally, NR_033928 upregulated the expression of glutaminase (GLS) by interacting with IGF2BP3/HUR complex to promote GLS mRNA stability. Increased glutamine metabolite, α-KG, upregulated NR_033928 expression by enhancing its promoter 5-hydroxymethylcytosine (hm5C) demethylation. In conclusion, our results revealed that NSUN2-methylated NR_033928 promoted GC progression and might be a potential prognostic and therapeutic target for GC.

Abstract Background The CRISPR/Cas12a and CRISPR/Cas13d systems are widely used for fundamental research and hold great potential for future clinical applications. However, the short half-life of guide RNAs (gRNAs), particularly free gRNAs without Cas nuclease binding, limits their editing efficiency and durability. Results Here, we engineer circular free gRNAs (cgRNAs) to increase their stability, and thus availability for Cas12a and Cas13d processing and loading, to boost editing. cgRNAs increases the efficiency of Cas12a-based transcription activators and genomic DNA cleavage by approximately 2.1- to 40.2-fold for single gene editing and 1.7- to 2.1-fold for multiplexed gene editing than their linear counterparts, without compromising specificity, across multiple sites and cell lines. Similarly, the RNA interference efficiency of Cas13d is increased by around 1.8-fold. In in vivo mouse liver, cgRNAs are more potent in activating gene expression and cleaving genomic DNA. Conclusions CgRNAs enable more efficient programmable DNA and RNA editing for Cas12a and Cas13d with broad applicability for fundamental research and gene therapy.

Abstract Keratinocyte hyperproliferation is a key pathogenic factor in psoriasis. However, the mechanisms that regulate keratinocyte hyperproliferation in this condition remain unclear. Here, we found that SLC35E1 was highly expressed in keratinocytes of patients with psoriasis and that Slc35e1 −/− mice displayed a less severe imiquimod (IMQ)-induced psoriasis-like phenotype than their wild-type siblings. In addition, SLC35E1 deficiency inhibited keratinocyte proliferation in both mice and cultured cells. On a molecular level, SLC35E1 was found to regulate zinc ion concentrations and subcellular localization, while zinc ion chelation reversed the IMQ-induced psoriatic phenotype in Slc35e1 −/− mice. Meanwhile, epidermal zinc ion levels were decreased in patients with psoriasis and zinc ion supplementation alleviated the psoriatic phenotype in an IMQ-induced mouse model of psoriasis. Our results indicated that SLC35E1 can promote keratinocyte proliferation by regulating zinc ion homeostasis and zinc ion supplementation has potential as a therapy for psoriasis.

Esophageal squamous cell carcinoma (ESCC) is an aggressive and fatal disease that is usually diagnosed when the chances for surgical intervention has been missed. Definitive concurrent chemoradiotherapy (dCRT) is the first choice of treatment for inoperable locally advanced esophageal squamous cell carcinoma (LA-ESCC). Nevertheless, the local recurrence rate for esophageal cancer patients undergoing dCRT remains high at 40-60%, with a 5-year overall survival rate of solely 10-30%. Immunotherapy in combination with dCRT is a promising treatment for inoperable LA-ESCC, for that improved long-term survival is expected. The present review provides a comprehensive overview of the evolutionary trajectory of dCRT for LA-ESCC, delineates notable relevant clinical studies, addresses unresolved concerns regarding the combination of dCRT with immunotherapy, and highlights promising directions for future research. When dCRT is combined with immunotherapy, the following aspects should be carefully explored in the future studies, including the optimal irradiation dose, segmentation scheme, radiotherapy technique, timing, sequence and duration of radiotherapy, and the selection of chemotherapeutic and immunologic drugs. In addition, further investigations on the mechanisms of how dCRT combined with immunotherapy exerts synergistic anti-tumor effects and molecular biomarkers ensuring precise screening of ESCC patients are needed.

Abstract The plasmid vector platform is the most commonly used vector for the expression of the versatile CRISPR-Cas technique and the promoter is a crucial element for the expression vector, thus profiling the impact of the promoters on CRISPR editors provides the basic information for the gene-editing toolkits and can be a guideline for its design. Herein, we made a parallel comparison among four commonly used promoters (CAG, ~ 1700 bp; EF1a core, ~ 210 bp; CMV, ~ 500 bp; and PGK, ~ 500 bp) in CRISPR-Cas12a system in mammalian cells to explore the impact of promoters on this powerful tool. We found that without badly damaging targeting specificity, the CAG promoter-driving Cas12a editor exhibited the most active (efficiency takes as 100%, specificity index = ~ 75%) in genomic cleavage, multiplex editing, transcriptional activation, and base editing, followed by promoter CMV (efficiency = 70 ~ 90% (vs CAG), specificity index = ~ 78%), and then EF1a core and PGK (both efficiency = 40–60%, vs CAG) but with higher specificity (specificity index = ~ 84% and ~ 82%, respectively). Therefore, CAG is recommended in the CRISPR-Cas12a system for the applications that need a robust editing activity but without size limitation, CMV mostly can be an alternative for CAG when requiring a smaller space, EF1a is similar to PGK with relatively high specificity, but has a smaller size, thus is more suitable for in vivo therapeutic applications. The data outlined the properties of the widely used promoters in the CRISPR-Cas12a system, which can be a guide for its applications and can be a useful resource for the gene-editing field.

Background: Pelareorep is an oncolytic virus that causes oncolytic effects in many solid tumors, and it has shown therapeutic benefits. However, few studies have compared pelareorep combined with chemotherapy to traditional chemotherapy alone in advanced solid tumors. Consequently, we intended to evaluate the effectiveness and safety of pelareorep plus chemotherapy in this paper.Methods: We searched four databases including PubMed, Embase, Cochrane Library and Web of Science comprehensively for studies comparing pelareorep combined with chemotherapy to chemotherapy alone in the treatment of advanced solid tumors. The outcomes measures were 1-year overall survival (OS), 2-year OS, 4-month progression-free survival (PFS), 1-year PFS, objective response rate (ORR), any-grade adverse events (any-grade AEs), and severe AEs (grade ≥ 3).Results: There were five studies involving 492 patients included in the study. Combination therapy did not significantly improve clinical outcomes in terms of 1-year OS [RR = 1.02, 95%CI = (0.82–1.25)], 2-year OS [RR = 1.00, 95%CI = (0.67–1.49)], 4-month PFS [RR = 1.00, 95%CI = (0.67–1.49)], 1-year PFS [RR = 0.79, 95%CI = (0.44–1.42)], and ORR [OR = 0.79, 95%CI = (0.49–1.27)] compared to chemotherapy alone, and the subgroup analysis of 2-year OS, 1-year PFS, and ORR based on countries and tumor sites showed similar results. In all grades, the incidence of AEs was greater with combination therapy, including fever [RR = 3.10, 95%CI = (1.48–6.52)], nausea [RR = 1.19, 95%CI = (1.02–1.38)], diarrhea [RR = 1.87, 95%CI = (1.39–2.52)], chills [RR = 4.14, 95%CI = (2.30–7.43)], headache [RR = 1.46, 95%CI = (1.02–2.09)], vomiting [RR = 1.38, 95%CI = (1.06–1.80)] and flu-like symptoms [RR = 4.18, 95%CI = (2.19–7.98)]. However, severe adverse events did not differ significantly between the two arms.Conclusion: Pelareorep addition to traditional chemotherapy did not lead to significant improvements in OS, PFS, or ORR in advanced solid tumor patients, but it did partially increase AEs in all grades, with no discernible differences in serious AEs. Therefore, the combination treatment is not recommended in patients with advanced solid tumors.Systematic Review Registration:https://www.crd.york.ac.uk/PROSPERO/display_record.php?RecordID=400841, identifier CRD42023400841

Abstract Background Profiling and comparing the performance of current widely used DNA targeting CRISPR systems provide the basic information for the gene-editing toolkit and can be a useful resource for this field. In the current study, we made a parallel comparison between the recently reported miniature Cas12f1 (Un1Cas12f1 and AsCas12f1) and the widely used Cas12a and Cas9 nucleases in mammalian cells. Results We found that as a CRISPRa activator, Un1Cas12f1 could induce gene expression with a comparable level to that of Cas12a and Cas9, while as a DNA cleavage editor, Cas12f1 exhibited similar properties to Cas12a, like high specificity and dominantly induced deletions over insertions, but with less activity. In contrast, wild-type SpCas9 showed the highest activity, lowest specificity, and induced balanced deletions and insertions. Thus, Cas12f1 is recommended for gene-activation-based applications, Cas12a is for therapy applications, and wild-type Cas9 is for in vitro and animal investigations. Conclusion The comparison provided the editing properties of the widely used DNA-targeting CRISPR systems in the gene-editing field.

Some cervical cancer patients show poor response to neoadjuvant chemotherapy. Although it has been demonstrated that macroautophagy/autophagy contributes to the poor chemosensitivity, the underlying mechanism has not yet been fully clarified. In this autophagy punctum, we discuss our recent findings about the mechanism of SOX6-induced autophagy and its potential significance in the platinum-based chemotherapy of cervical cancer.

Abandoned cropland is a great waste of land resources, which not only affects the stability of food production, but also wastes limited cropland resources. China has large areas of abandoned cropland, and accurately extracting the large-scale abandoned cropland is a prerequisite for the government to implement reclamation policies and estimate the food production potential of abandoned cropland. China has implemented the Grain for Green Project (GGP) since 1980, but most research has not considered its impact on the extraction of abandoned cropland. Therefore, a method was proposed to identify abandoned cropland excluding the regions of GGP, and to analyze the spatio-temporal characteristics and potential of abandoned cropland in China from 1991 to 2018. GGP potential and food production potential of abandoned cropland were discussed. The results showed that 26.47 million hectares of cropland has been abandoned in the past 28 years (including cropland that has been reclaimed). Under the positive influence of the government’s food security policy, the abandoned cropland has decreased in recent years, but the total area is still high. The abandonment mainly occurred in the Jiangnan Hills, the Hilly Regions of Fujian and Guangdong Province, the North China Mountains, and Eastern Inner Mongolia. The higher the elevation and the steeper the slope, the higher the abandonment rate. In addition, the urban-rural income gap significantly affected the abandonment rate. This study drew the spatial and temporal distribution map of abandoned cropland in China with a spatial resolution of 30 m considering GGP. It provides an important basis for formulating reclamation and GGP policy.

Abstract Background The CRISPR-Cas12a (formerly Cpf1) system is a versatile gene-editing tool with properties distinct from the broadly used Cas9 system. Features such as recognition of T-rich protospacer-adjacent motif (PAM) and generation of sticky breaks, as well as amenability for multiplex editing in a single crRNA and lower off-target nuclease activity, broaden the targeting scope of available tools and enable more accurate genome editing. However, the widespread use of the nuclease for gene editing, especially in clinical applications, is hindered by insufficient activity and specificity despite previous efforts to improve the system. Currently reported Cas12a variants achieve high activity with a compromise of specificity. Here, we used structure-guided protein engineering to improve both editing efficiency and targeting accuracy of Acidaminococcus sp. Cas12a (AsCas12a) and Lachnospiraceae bacterium Cas12a (LbCas12a). Results We created new AsCas12a variant termed “AsCas12a-Plus” with increased activity (1.5~2.0-fold improvement) and specificity (reducing off-targets from 29 to 23 and specificity index increased from 92% to 94% with 33 sgRNAs), and this property was retained in multiplex editing and transcriptional activation. When used to disrupt the oncogenic BRAFV600E mutant, AsCas12a-Plus showed less off-target activity while maintaining comparable editing efficiency and BRAFV600E cancer cell killing. By introducing the corresponding substitutions into LbCas12a, we also generated LbCas12a-Plus (activity improved ~1.1-fold and off-targets decreased from 20 to 12 while specificity index increased from 78% to 89% with 15 sgRNAs), suggesting this strategy may be generally applicable across Cas12a orthologs. We compared Cas12a-Plus, other variants described in this study, and the reported enCas12a-HF, enCas12a, and Cas12a-ultra, and found that Cas12a-Plus outperformed other variants with a good balance for enhanced activity and improved specificity. Conclusions Our discoveries provide alternative AsCas12a and LbCas12a variants with high specificity and activity, which expand the gene-editing toolbox and can be more suitable for clinical applications.

Abstract There are nearly 40% of cervical cancer patients showing poor response to neoadjuvant chemotherapy that can be induced by autophagy, however, the underlying mechanism has not yet been fully clarified. We previously found that Sex-determining region of Y-related high-mobility-group box 6 (SOX6), a tumor suppressor gene or oncogene in several cancers, could induce autophagy in cervical cancer. Accordingly, this study aims to investigate the mechanism of SOX6-induced autophagy and its potential significance in the platinum-based chemotherapy of cervical cancer. Firstly, we found that SOX6 could promote autophagy in cervical cancer cells depending on its HMG domain. Mitogen-activated protein kinase kinase kinase kinase-4 (MAP4K4) gene was identified as the direct target gene of SOX6, which was transcriptionally upregulated by binding the HMG domain of SOX6 protein to its double-binding sites within MAP4K4 gene promoter. MAP4K4 mediated the SOX6-induced autophagy through inhibiting PI3K-Akt-mTOR pathway and activating MAPK/ERK pathway. Further, the sensitivity of cervical cancer cells to cisplatin chemotherapy could be reduced by the SOX6-induced autophagy in vitro and in vivo, while such a phenomenon could be turned over by autophagy-specific inhibitor and MAP4K4 inhibitor, respectively. Moreover, cisplatin itself could promote the expression of endogenous SOX6 and subsequently the MAP4K4-mediated autophagy in cervical cancer cells, which might in turn reduce the sensitivity of these cells to cisplatin treatment. These findings uncovered the underlying mechanism and potential significance of SOX6-induced autophagy, and shed new light on the usage of MAP4K4 inhibitor or autophagy-specific inhibitor for sensitizing cervical cancer cells to the platinum-based chemotherapy.

In recent years, the vegetation cover in the Selinco region of the Qinghai-Tibet Plateau has undergone significant changes due to the influence of global warming and intensified human activity. Consequently, comprehending the distribution and change patterns of vegetation in this area has become a crucial scientific concern. To address this concern, the present study employed MODIS-NDVI and elevation data, integrating methodologies such as trend analysis, Hurst exponent analysis, and sequential cluster analysis to explore vegetation cover changes over the past 21 years and predict future trends, while examining their correlation with topographic factors. The study findings indicate a fluctuating upward trend in vegetation cover, with a notable decrease in 2015. Spatially, the overall fractional vegetation cover (FVC) in the study area showed a basic stability with a percentage of 78%. The analysis of future trends in vegetation cover revealed that the majority of areas (68.26%) exhibited an uncertain trend, followed by stable regions at 15.78%. The proportion of areas showing an increase and decrease in vegetation cover accounted for only 9.63% and 5.61%, respectively. Elevation and slope significantly influence vegetation cover, with a trend of decreasing vegetation cover as elevation increases, followed by an increase, and then another decrease. Likewise, as the slope increases, initially, there is a rise in vegetation cover, followed by a subsequent decline. Notably, significant abrupt changes in vegetation cover are observed within the 4800 m elevation band and the 4° slope band in the Selinco region. Moreover, aspect has no significant effect on vegetation cover. These findings offer comprehensive insights into the spatial and temporal variations of vegetation cover in the Selinco region and their association with topographic factors, thus serving as a crucial reference for future research.

Hongxin Huang and Yongfei Hu et al. develop Tag-seq, a streamlined sequencing method to examine off-target editing by CRISPR-Cas systems. The authors validate Tag-seq in multiple cell lines, demonstrating its broad utility and flexibility in identifying off-target genome editing.

To obtain a Johnson–Cook model of 15-5 PH steel formed by selective laser melting (SLM), and to determine the difference between the forging process, in this work, mechanical testing, penetration testing and numerical simulations were used to study 15-5 PH steel formed by SLM and forging. Finally, the Johnson–Cook model parameters and failure parameters of the 15-5 PH steel formed by SLM and forging were obtained. We found that the SLM process was beneficial for refining the grain size of 15-5 PH steel and for improving the mechanical properties of 15-5 PH steel, where the yield strength of its specimens increased by 13.1% compared with the forged specimens. The error between the numerical simulations and penetration tests was less than 10%, which verified the validity of the numerical model parameters. It was also found that the penetration ability and abrasion resistance of the SLM-shaped projectiles were slightly superior to those of the forged projectiles.

Abstract The use of immunotherapy has achieved great advances in the treatment of cancer. Macrophages play a pivotal role in the immune defense system, serving both as phagocytes (removal of pathogens and cancer cells) and as antigen‐presenting cells (activation of T cells). However, research regarding tumor immunotherapy is mainly focused on the adaptive immune system. The usefulness of innate immune cells (eg, macrophages) in the treatment of cancer has not been extensively investigated. Recent advances in synthetic biology and the increasing understanding of the cluster of differentiation 47/signal regulatory protein alpha (CD47/SIRPɑ) axis may provide new opportunities for the clinical application of engineered macrophages. The CD47/SIRPɑ axis is a major known pathway, repressing phagocytosis and activation of macrophages. In this article, we summarize the currently available evidence regarding the CD47/SIRPɑ axis, and immunotherapies based on blockage. In addition, we propose cell therapy strategies based on macrophage engineering.

Rock penetration is an inevitable problem in the study of drilling and projectile penetration. The penetration resistance of red sandstone and limestone was investigated at projectile speeds ranging from 600–1200 m/s. The damage characteristics of these rocks were studied via experimental tests and numerical simulations. The damage condition of the target surface, internal damage state and crack distribution were obtained. It was concluded that the maximum error of the numerical simulation and experimental results was not more than 10%. The penetration resistance of limestone was approximately 23.8% stronger than that of red sandstone. However, the energy absorption effect of limestone was weaker than that of red sandstone, and large cracks can be more easily formed. The compaction area of red sandstone was softer, with obvious crack compaction in the crater area, and particle detachment can more easily occur. Red sandstone was more sensitive to the impact angle of the projectile. With oblique penetration, the projectile was more likely to deflect inside the red sandstone target.

Simple systems in which strong coupling of different excitations can be easily realized are highly important, not only for fundamental research but also for practical applications. Here, we proposed a T-shaped gold nanorod (GNR) dimer composed of a long GNR and a short GNR perpendicular to each other and revealed that the dark quadrupole mode of the long GNR can be activated by utilizing the dipole mode excited in the short GNR. It was found that the strong coupling between the dipole and quadrupole modes can be achieved by exciting the T-shaped GNR dimer with a plane wave. Then, we demonstrated the realization of strong dipole–quadrupole–exciton coupling by placing a T-shaped GNR on a tungsten disulfide (WS2) monolayer, which leads to a Rabi splitting as large as ~299 meV. It was confirmed that the simulation results can be well fitted by using a Hamiltonian based on the coupled harmonic oscillator model and the coupling strengths for dipole–quadrupole, dipole–exciton and quadrupole–exciton can be extracted from the fitting results. Our findings open new horizons for realizing strong plasmon–exciton coupling in simple systems and pave the way for constructing novel plasmonic devices for practical applications.

Background/Objective: Tanshinol is the main active component of Salvia miltiorrhiza Bunge, a significant Traditional Chinese Medicine used to treat cardiovascular disease. We have shown that tanshinol exerts an antiosteoporostic effect via the enhancement of bone formation in vivo and in vitro. However, the mechanism remains unclear. Based on the polyphenol group in the structure of tanshinol, we speculate the protective action on skeletal tissue is related to antioxidative capacity. Our in vitro evidence indicated that tanshinol stimulated osteoblastic differentiation by its antioxidaive capacity. In this study, we aim to further confirm the effect of tanshinol on bone formation and the underlying mechanism in zebrafish in vivo. Methods: We used a Danio rerio (zebrafish) model, which has a bone formation process similar to humans, and evaluated the relationship between the dose and the effect of tanshinol on bone formation determined using alizarin red S staining or fluorescence intensity analysis in normal and glucocorticoid (GC)-induced inhibition of an osteogenesis model using wild-type zebrafish and cortical bone transgenic zebrafish tg(sp7:egfp). The expression of osteoblast-specific genes and reactive oxygen species (ROS) were tested. Results: Our data showed that dexamethasone exerts a series of consequences, including the inhibition of bone formation, decrease of bone mass, downregulation of expression of osteoblast-specific genes (runx2a, ALP, osteocalcin, and sp7), as well as the accumulation of ROS generation and decreased capacity of antioxidants. Tanshinol showed a protective effect on promoting bone formation and bone mass both in wild-type larval zebrafish and transgenic zebrafish. Furthermore, tanshinol attenuated the inhibition of osteogenesis elicited by oxidative stress in the zebrafish exposed to dexamethasone. Conclusion: The present findings suggest that tanshinol prevented decreased osteogenesis in GC-treated larval zebrafish via scavenging ROS and stimulated the expression of osteoblast-specific genes. Tanshinol treatment may be developed as a novel therapeutic approach under recent recognised conditions of GC-induced osteoporosis.

A generalized contour-sum method has been proposed to measure the topological charge (TC) of an optical vortex (OV) beam using a Shack−Hartmann wavefront sensor (SH-WFS). Moreover, a recent study extended it to be workable for measuring an aberrated OV beam. However, when the OV beam suffers from severe distortion, the closed path for circulation calculation becomes crucial. In this paper, we evaluate the performance of five closed path determination methods, including watershed transformation, maximum average-intensity circle extraction, a combination of watershed transformation and maximum average-intensity circle extraction, and perfectly round circles assignation. In the experiments, we used a phase-only spatial light modulator to generate OV beams and aberrations, while an SH-WFS was used to measure the intensity profile and phase slopes. The results show that when determining the TC values of distorted donut-shaped OV beams, the watershed-transformed maximum average-intensity circle method performed the best, and the maximum average-intensity circle method and the watershed transformation method came second and third, while the worst was the perfect circles assignation method. The discussions that explain our experimental results are also given.

Human Papillomavirus (HPV) 16 infection is considered as one of the significant causes of human cervical cancer. The expression of the viral oncogenes like E6 and E7 play an important role in the development of the cancer. MiR-122 has been reported to exhibit a strong relationship with hepatitis viruses and take part in several tumor development, while the effects of miR-122 on HPV infection and the HPV viral oncogenes expression still remain unexplored. In this study, using RNAhybrid software, the potential binding sites between miR-122 and HPV16 E6 and E7 mRNAs were identified. Over and loss of miR-122 function showed that miR-122 could directly bind with HPV16 E6 mRNA and significantly inhibit its expression in SiHa cells, which was further confirmed by constructing the miR-122-E6-mu to eliminate the miR-122 binding effects with E6. The increase of the expression of type I interferon (IFN) and its classical effective molecules and the phosphorylation of signal transducers and activators of transcription (STAT1) protein indicated that miR-122 might enhance type I interferon in cervical carcinoma cells, which explained the significant reduction of HPV16 E7 and E6*I mRNA expression. This might be due to the binding between miR-122 and suppressor of cytokine signaling 1 (SOCS1) mRNA, which is the suppressor of interferon signaling pathway. Moreover, it was identified that the miR-122 binding position was nt359-nt375 in SOCS1 mRNA. Taken together, this study indicated that HPV16 could be effectively inhibited by miR-122 through both direct binding with E6 mRNA and promoting SOCS1-dependent IFN signaling pathway. Thus, miR-122 may serve as a new therapeutic option for inhibiting HPV infection.

BACKGROUND: The risk of fatal adverse events (FAEs) due to bevacizumab-based chemotherapy has not been well described; we carried out an updated meta-analysis regarding this issue. METHODS: An electronic search of Medline, Embase and The Cochrane Central Register of Controlled Trials was conducted to investigate the effects of randomized controlled trials on bevacizumab treatment on cancer patients. Random or fixed-effect meta-analytical models were used to evaluate the risk ratio (RR) of FAEs due to the use of bevacizumab. RESULTS: Thirty-four trials were included. Allocation to bevacizumab therapy significantly increased the risk of FAEs; the RR was 1.29 (95% CI:1.05-1.57). This association varied significantly with tumor types (P=0.002) and chemotherapeutic agents (P=0.005) but not with bevacizumab dose (P=0.90). Increased risk was seen in patients with non-small cell lung cancer, pancreatic cancer, prostate cancer, and ovarian cancer. However, FAEs were lower in breast cancer patients treated with bevacizumab. In addition, bevacizumab was associated with an increased risk of FAEs in patients who received concomitant agents of taxanes and/or platinum. CONCLUSION: Compared with chemotherapy alone, the addition of bevacizumab was associated with an increased risk of FAEs among patients with special tumor types, particularly when combined with chemotherapeutic agents such as platinum.