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Objective To investigate the application effect of transcutaneous electrical acupoint stimulation (TEAS) combined with propofol in painless gastrointestinal endoscopy. Methods A total of 105 patients who underwent gastrointestinal endoscopy were randomly divided into propofol+fentanyl group (P+F group), propofol+TEAS group (P+T group), and propofol+placebo-TEAS group (P+P group), with 35 patients in each group. All patients were induced to deep sedation by the anesthesiologist using propofol before gastroscopy; the patients in the P+F group were given intravenously injected fentanyl 50 μg before gastroscopy, those in the P+T group received placement of electrodes at the acupoints at 30 minutes before examination and were given electrical stimulation till the end of the examination, and those in the P+P group received placement of electrodes at the acupoints alone without electrical stimulation. The patients were recorded in terms of pain score and the amount of propofol used du-ring examination, the time to recovery from anesthesia after examination, and the incidence rate of adverse reactions during or after examination, as well as the degree of satisfaction among patients and physicians. Results Compared with the P+P group, the P+F and P+T groups had a significant reduction in pain score (H=23.16,P0.05), while the P+F group had significantly higher degrees of vertigo, nausea, and vomiting than the other two groups (H=40.97-44.79,P

Abstract Background The global prevalence of VCI has increased steadily in recent years, but diagnostic biomarkers for VCI in patients with non-disabling ischemic cerebrovascular incidents (NICE) remain indefinite. The primary objective of this research was to investigate the relationship between peripheral serological markers, white matter damage, and cognitive function in individuals with NICE. Methods We collected clinical data, demographic information, and medical history from 257 patients with NICE. Using the MoCA upon admission, patients were categorized into either normal cognitive function (NCF) or VCI groups. Furthermore, they were classified as having mild white matter hyperintensity (mWMH) or severe WMH based on Fazekas scores. We then compared the levels of serological markers between the cognitive function groups and the WMH groups. Results Among 257 patients with NICE, 165 were male and 92 were female. Lymphocyte count (OR = 0.448, P

Abstract Background Tobacco bacterial wilt (TBW) caused by Ralstonia solanacearum is the most serious soil-borne disease of tobacco that significantly reduces crop yield. However, the limited availability of resistance in tobacco hinders breeding efforts for this disease. Results In this study, we conducted hydroponic experiments for the root expression profiles of D101 (resistant) and Honghuadajinyuan (susceptible) cultivars in response to BW infection at 0 h, 6 h, 1 d, 3 d, and 7d to explore the defense mechanisms of BW resistance in tobacco. As a result, 20,711 and 16,663 (total: 23,568) differentially expressed genes (DEGs) were identified in the resistant and susceptible cultivars, respectively. In brief, at 6 h, 1 d, 3 d, and 7 d, the resistant cultivar showed upregulation of 1553, 1124, 2583, and 7512 genes, while the susceptible cultivar showed downregulation of 1213, 1295, 813, and 7735 genes. Similarly, across these time points, the resistant cultivar had downregulation of 1034, 749, 1686, and 11,086 genes, whereas the susceptible cultivar had upregulation of 1953, 1790, 2334, and 6380 genes. The resistant cultivar had more up-regulated genes at 3 d and 7 d than the susceptible cultivar, indicating that the resistant cultivar has a more robust defense response against the pathogen. The GO and KEGG enrichment analysis showed that these genes are involved in responses to oxidative stress, plant–pathogen interactions, cell walls, glutathione and phenylalanine metabolism, and plant hormone signal transduction. Among the DEGs, 239 potential candidate genes were detected, including 49 phenylpropane/flavonoids pathway-associated, 45 glutathione metabolic pathway-associated, 47 WRKY, 48 ERFs, eight ARFs, 26 pathogenesis-related genes (PRs), and 14 short-chain dehydrogenase/reductase genes. In addition, two highly expressed novel genes (MSTRG.61386-R1B-17 and MSTRG.61568) encoding nucleotide-binding site leucine-rich repeat (NBS-LRR) proteins were identified in both cultivars at 7 d. Conclusions This study revealed significant enrichment of DEGs in GO and KEGG terms linked to glutathione, flavonoids, and phenylpropane pathways, indicating the potential role of glutathione and flavonoids in early BW resistance in tobacco roots. These findings offer fundamental insight for further exploration of the genetic architecture and molecular mechanisms of BW resistance in tobacco and solanaceous plants at the molecular level.

In our previous research, we found that trichokonins’ (TKs) employment improved the thermotolerance of the Lanzhou lily, a renowned edible crop species endemic to China that is relatively susceptible to high temperatures (HTs). Here, a novel Lanzhou lily GRAS gene, LzSCL9, was identified to respond to heat stress (HS) and HS+TKs treatment based on transcriptome and RT-qPCR analysis. TKs could improve the upregulation of LzSCL9 during long-term HS. The expression profile of LzSCL9 in response to HS with or without TKs treatment showed a significant positive correlation with LzHsfA2a-1, which was previously identified as a key regulator in TKs’ conferred resilience to HT. More importantly, overexpression of LzSCL9 in the lily enhanced its tolerance to HTs and silencing LzSCL9 in the lily reduced heat resistance. Taken together, this study identified the positive role of LzSCL9 in TK-induced thermotolerance, thereby preliminarily establishing a molecular mechanism on TKs regulating the thermostability of the Lanzhou lily and providing a new candidate regulator for plant heat-resistant breeding.

Mantle cell lymphoma (MCL) is a mature B-cell lymphoma associated with cyclin D family rearrangements and typically expresses CD5 and cyclin D1. Epstein–Barr virus- (EBV-) positive MCL is rare, and the role of EBV infection and its transformation in MCL remains unclear. We present a case of CD5-negative classic MCL that progressed to an EBV + pleomorphic MCL six years after the initial diagnosis. Molecular studies confirmed the same clonal origin. To the best of our knowledge, the EBV-positive transformation of CD5-negative MCL into a pleomorphic variant has rarely been reported, and its recognition is important for the diagnosis and the management of patients with MCL.

Acetohydroxyacid synthase (AHAS) is a key enzyme in the first step of the branched-chain amino acid synthesis pathway, and the production of acetohydroxybutyrate from one molecule of 2-ketobutyric acid and one molecule of pyruvate. AHAS is inhibited by feedback from L-valine, L-leucine, and L-isoleucine, and the expression of ilvBN, the gene encoding AHAS, is regulated by all three branched-chain amino acids. A change in amino acids 20–22 on the regulatory subunit (M13 mutation) removes the feedback inhibition by valine. We cloned the gene encoding AHAS (ilvBN) into a vector and then transfected it into Escherichia coli BL21 for expression with targeted changes in amino acids 20–22 on the regulatory subunit, and then determined the activity of the mutated AHAS and its inhibitory effects on valine, isoleucine, and leucine. The enzyme containing the M13 mutation was feedback resistant to all three amino acids. Previous studies have suggested that the binding sites for the three branched-chain amino acids may be at the same variable center. We investigated the enzymatic properties of wild-type and mutant AHAS, modeled their crystal structures, and resolved the mechanism of feedback inhibition induced by mutant M13, which will be useful for continuing the modification of AHAS and the design of broad-spectrum herbicides.

Fired bricks for building were prepared from the tailings of vanadium extraction from stone coal. The influences of fineness and addition of tailing on the water absorption and compressive strength of samples were investigated. The mechanism of tailoring the phase compositions and microstructures of samples by tailing addition was analyzed by XRD and SEM. The freeze-thaw resistance and utilization safety of the products were tested. The results showed increasing the tailing fineness enhanced the microstructure densification and compressive strength, which firstly increased and then decreased after increasing tailing addition. B3 sample (tailings∶Clay∶fly ash = 30∶30∶40) fired at 1150 ℃ had the compressive strength of ≥35 MPa and the water absorption of ≤ 13%. The comprehensive properties meet the requirements of the Mu30 standard in the《sintered ordinary bricks》(GB/T 5101-2003). XRD and SEM analysis showed that adding 15%~30% tailing was beneficial to the formation of glass phase and promote the production of anorthite as well as anorthosite. Grain and liquid phase are cemented with each other, thus endowing the products with high compressive strength, microstructure densification, and good freeze-thaw resistance.

Summary: CircRNAs play multiple roles in a variety of cellular processes. We found that Circ-CDYL is highly enriched in early HCC plasma exosomes. Moreover, EpCAM+ HCC cells and exosomes had significant Circ-CDYL levels. We postulated that Circ-CDYL-enriched and EpCAM-positive exosomes would function as liver tumor-initiating exosomes (LTi-Exos). As predicted, intercellular transfer of LTi-Exos activates the HDGF-PI3K-AKT-mTOR and HIF1AN-NOTCH2 axes in recipient cells, promoting malignancy. Upstream, we found that the N6-methyladenosine (m6A) modification of Circ-CDYL exerted its action in HCC cells through a dual mechanism. First, it stimulated back-splicing processes via YTHDC1 to promote Circ-CDYL biogenesis. Second, it facilitates the active sorting of Circ-CDYL into exosomes via hnRNPA2/B1. Clinically, the combination of LTi-Exos and plasma alpha-fetoprotein (AFP) provides a promising early diagnostic biomarker for HCC with an AUC of 0.896. This study highlights the effect and mechanism by which m6A modification promotes hepatocarcinogenesis via modulation of the tumor microenvironment by LTi-Exos.

BackgroundMajor depression disorder (MDD) is a devastating neuropsychiatric disease, and one of the leading causes of suicide. Ferroptosis, an iron-dependent form of regulated cell death, plays a pivotal role in numerous diseases. The study aimed to construct and validate a gene signature for diagnosing MDD based on ferroptosis-related genes (FRGs) and further explore the biological functions of these genes in MDD.MethodsThe datasets were downloaded from the Gene Expression Omnibus (GEO) database and FRGs were obtained from the FerrDb database and other literatures. Least absolute shrinkage and selection operator (LASSO) regression and stepwise logistic regression were performed to develop a gene signature. Receiver operating characteristic (ROC) curves were utilized to assess the diagnostic power of the signature. Gene ontology (GO) enrichment analysis was used to explore the biological roles of these diagnostic genes, and single sample gene set enrichment analysis (ssGSEA) algorithm was used to evaluate immune infiltration in MDD. Animal model of depression was constructed to validate the expression of the key genes.ResultsEleven differentially expressed FRGs were identified in MDD patients compared with healthy controls. A signature of three FRGs (ALOX15B, RPLP0, and HP) was constructed for diagnosis of MDD. Afterwards, ROC analysis confirmed the signature’s discriminative capacity (AUC = 0.783, 95% CI = 0.719–0.848). GO enrichment analysis revealed that the differentially expressed genes (DEGs) related to these three FRGs were mainly involved in immune response. Furthermore, spearman correlation analysis demonstrated that these three FRGs were associated with infiltrating immune cells. ALOX15B and HP were significantly upregulated and RPLP0 was significantly downregulated in peripheral blood of the lipopolysaccharide (LPS)-induced depressive model.ConclusionOur results suggest that the novel FRG signature had a good diagnostic performance for MDD, and these three FRGs correlated with immune infiltration in MDD.

[Objective] The spatial change characteristics and spatial aggregation characteristics of important ecological protection areas in the Poyang Lake urban agglomeration were analyzed in order to provide a basis for delineating "three districts and three lines" and rational planning of territorial space, and to provide a theoretical reference for promoting the coordinated development between high-quality economic development and ecological protection in the Great Lake Basin. [Methods] The study area was the urban agglomeration around Poyang Lake. Land use data for 2000, 2010, and 2020 were used with the InVEST model, the revised universal soil loss equation (RUSLE), and a GIS spatial analysis method to select types of ecological services (such as water conservation, soil conservation, biodiversity conservation, water purification, soil erosion sensitivity, and water environment sensitivity) to identify the important areas of ecological protection in the study area, and to analyze their spatial variation characteristics. [Results] ① In 2000, 2010, and 2020, the areas of important ecological protection in the urban agglomeration around Poyang Lake were 6.34×104, 6.35×104, and 6.28×104 km2, respectively. The land use types were dominated by forest land and water areas, showing the distribution characteristics of "high on both sides and low in the middle" . ② At a grid scale of 2 km×2 km, the importance of ecological protection in the urban agglomeration around Poyang Lake was significantly clustered in space, and the high and high clustering areas were mainly located in the eastern and western mountainous areas with fewer anthropogenic activities and good ecological protection. Low and low agglomeration areas were mainly located in the plain area around the Poyang Lake (where human activities are intensive), and some functions and structures of the ecosystem itself were damaged during economic development. ③ From 2000 to 2020, the changing and stable areas of ecological protection importance were located in the Huaiyu mountain area and the Poyang Lake Plain, and the enhanced areas were located in Fuliang County, Wuyuan County, and the Poyang Lake District; and the weakened areas were located in Yichun City, Pingxiang City, and Yingtan City. [Conclusion] For extremely important and highly important areas, stabilizing forest coverage and improving forest quality is the first task. Secondly, measures should be taken to effectively promote wetland restoration and the construction of nature reserves in order to curb habitat degradation. For medium-important, moderately important, and less important areas, the relationship between high-quality economic development and ecological protection should be properly handled. Especially in areas with intensive human activities, urban green space coverage should be increased, and ecological buffer zones should be established around cities to alleviate the pressure of ecological space survival.

Graves' orbitopathy (GO) is an autoimmune disease that involves complex immune systems. The mainstays of clinical management for this disease are surgery, targeted drugs therapy, and no-targeted drugs drug therapy. targeted drugs can improve therapeutic efficacy and enhance the quality of life for GO patients. However, as a second-line treatment for GO, targeted drugs such as tocilizumab and rituximab have very limited therapeutic effects and may be accompanied by side effects. The introduction of Teprotumumab, which targets IGF-IR, has made significant progress in the clinical management of GO. The pathophysiology of GO still remains uncertain as it involves a variety of immune cells and fibroblast interactions as well as immune responses to relevant disease targets of action. Therfore, learning more about immune response feedback pathways and potential targets of action will assist in the treatment of GO. In this discussion, we explore the pathogenesis of GO and relevant work, and highlight four potential targets for GO: Interleukin-23 receptor (IL-23 R), Leptin receptor (LepR), Orbital fibroblast activating factors, and Plasminogen activator inhibitor-1 (PAI-1). A deeper understanding of the pathogenesis of GO and the role of potential target signaling pathways is crucial for effective treatment of this disease.

Background The turnover intention level of general practitioners (GPs) in primary care in China is relatively high, and identification of its associated factors will offer insights into the curbing of outflow of health talents in primary care. The targeted admission medical graduates (TAMG) who completed the "5+3" standardized general residency program (5-year undergraduate medical education plus 3-year standardized residency training) have gradually entered primary care to fulfill their preadmission commitments, but research on their turnover intention after the tenure is few. Objective To explore the turnover intention and associated factors in Guizhou's TAMG trained with the "5+3" standardized general residency program after fulfilling their preadmission commitments in primary care, providing a basis for improving the policies about retaining the in-service health talents and developing the general practice workforce. Methods Participants were TAMG who were working in primary hospitals of Guizhou for fulfilling their preadmission commitments after finishing the "5+3" standardized general residency program at the end of 2020 (with a starting time for the 3-year standardized residency training of 2015—2017) . An online questionnaire survey was conducted with them from January 20 to February 10, 2021 for understanding their general information, self-assessed job satisfaction, turnover intention and planned career development after the expiration of services. Three hundred and eleven (89.6%) of the 347 cases who returned responsive questionnaires were selected as the final participants. Univariate and stepwise multiple linear regression analyses were used to explore the factors related to turnover intention. Results The average turnover intention score for the respondents was (3.98±0.98) points. The prevalence of having turnover intention was 73.6% (229/311) . The level of turnover intention differed obviously by sex, hospital location, and daily workload (P

ObjectiveDiagnosis classification and risk stratification are crucial in the prognosis prediction and treatment selection of acute myeloid leukemia (AML). Here, we used a database of 536 AML patients to compare the 4th and 5th WHO classifications and the 2017 and 2022 versions of ELN guidance.MethodsAML patients were classified according to the 4th and 5th WHO classifications, as well as the 2017 and 2022 versions of the European LeukemiaNet (ELN) guidance. Kaplan–Meier curves with log-rank tests were used for survival analysis.ResultsThe biggest change was that 25 (5.2%), 8 (1.6%), and 1 (0.2%) patients in the AML, not otherwise specified (NOS) group according to the 4th WHO classification, were re-classified into the AML-MR (myelodysplasia-related), KMT2A rearrangement, and NUP98 rearrangement subgroups based on the 5th WHO classification. Referring to the ELN guidance, 16 patients in the favorable group, six patients in the adverse group, and 13 patients in the intermediate group based on the 2017 ELN guidance were re-classified to the intermediate and adverse groups based on the 2022 ELN guidance. Regrettably, the Kaplan–Meier curves showed that the survival of intermediate and adverse groups could not be distinguished well according to either the 2017 or 2022 ELN guidance. To this end, we constructed a risk model for Chinese AML patients, in which the clinical information (age and gender), gene mutations (NPM1, RUNX1, SH2B3, and TP53), and fusions (CBFB::MYH11 and RUNX1::RUNX1T1) were included, and our model could help divide the patients into favorable, intermediate, and adverse groups.ConclusionThese results affirmed the clinical value of both WHO and ELN, but a more suitable prognosis model should be established in Chinese cohorts, such as the models we proposed.

Mast cell disorders range from benign proliferations to systemic diseases that cause anaphylaxis and other diverse symptoms to mast cell neoplasms with varied clinical outcomes. Mastocytosis is the pathologic process of the accumulation of abnormal mast cells in different organs, mostly driven by KIT mutations, and can present as cutaneous mastocytosis, systemic mastocytosis (SM), and mast cell sarcoma. The WHO 5th edition classification divides systemic mastocytosis into bone marrow mastocytosis, indolent systemic mastocytosis, smoldering systemic mastocytosis, aggressive systemic mastocytosis, systemic mastocytosis with an associated hematologic neoplasm, and mast cell leukemia. The new ICC classifies SM slightly differently. The diagnosis of SM requires the integration of bone marrow morphologic, immunophenotypic, and molecular findings, as well as clinical signs and symptoms. Moreover, understanding the wide range of clinical presentations for patients with mast cell disorders is necessary for accurate and timely diagnosis. This review provides an updated overview of mast cell disorders, with a special emphasis on SM, including the latest approaches to diagnosis, prognostic stratification, and management of this rare disease.

Chronic myeloid leukemia (CML) and chronic myelomonocytic leukemia (CMML) are two common myeloid neoplasms with overlapping morphologic features. We report a patient initially diagnosed with CML and treated with Tyrosine kinase inhibitor (TKI) but who then developed persistent monocytosis and worsening thrombocytopenia one year later. Repeat bone marrow biopsies only showed CML at the molecular level. However, markedly hypercellular bone marrow, megakaryocytic dysplasia, and SRSF2, TET2, and RUNX1 mutations by NextGen sequencing pointed to a diagnosis of CMML. For CML patients with persistent monocytosis and cytopenia, a mutational profile by NGS is helpful to exclude or identify the coexisting CMML.

The interferon (IFN) response is the first line of defense against viral invasion and thus plays a central role in the regulation of the immune response. IFN-epsilon (IFN-ε) is a newly discovered type I IFN that does not require viral induction, unlike other type I IFNs. IFN-ε is constitutively expressed in epithelial cells and plays an important role in mucosal immunity. In this study, we evaluated the biological activity of the mink-IFN (MiIFN)-ε gene in prokaryotic cells. Quantitative reverse transcription-polymerase chain reaction (qRT-PCR) was used to evaluate IFN-ε expression in different mink tissues. MiIFN-ε was highly expressed in brain, lung, tracheal, kidney, intestinal, bladder, ovarian, and testis tissues. There was no significant difference in MiIFN-ε expression between female and male minks, except in the reproductive system. Expression of the small ubiquitin-like modifier (SUMO3)-MiIFN-ε fusion gene was induced by isopropylβ-d-thiogalactoside, and MiIFN-ε was collected after SUMO-specific protease digestion. We tested the antiviral activity of MiIFN-ε against vesicular stomatitis virus (VSV) in epithelial cells of feline kidney 81 (F81). We used qRT-PCR to analyze the expression of several IFN-stimulated genes (ISGs), including ISG15, 2′-5′ oligoadenylate synthetase (2′-5′OAS1), and myxovirus resistance protein 1 (Mx1). Recombinant IFN-ε induced high ISG expression in F81 cells. Compared with those in the cell control group, expressions of ISG15, Mx1, and 2′-5′ OAS1 in the VSV-GFP control, IFN-ε, and MiIFN-ε-inhibited VSV-GFP groups were significantly increased. Compared with those in the VSV-GFP control group, expressions of ISG15 and 2′-5′ OAS1 in the IFN-ε and MiIFN-ε-inhibited VSV-GFP groups were significantly increased, and the differences were highly significant (p < 0.0001). IFN-ε played an indirect antiviral role. These findings lay the foundation for detailed investigation of IFN-ε in the future.

Canine distemper (CD), caused by canine distemper virus (CDV), is a highly contagious and lethal disease in domestic and wild carnivores. Although CDV live-attenuated vaccines have reduced the incidence of CD worldwide, low levels of protection are achieved in the presence of maternal antibodies in juvenile animals. Moreover, live-attenuated CDV vaccines may retain residual virulence in highly susceptible species and cause disease. Here, we generated several CDV DNA vaccine candidates based on the biscistronic vector (pIRES) co-expressing virus wild-type or codon-optimized hemagglutinin (H) and nucleocapsid (N) or ferret interferon (IFN)-γ, as a molecular adjuvant, respectively. Apparently, ferret (Mustela putorius furo)-specific codon optimization increased the expression of CDV H and N proteins. A ferret model of CDV was used to evaluate the protective immune response of the DNA vaccines. The results of the vaccinated ferrets showed that the DNA vaccine co-expressing the genes of codon-optimized H and ferret IFN-γ (poptiH-IRES-IFN) elicited the highest anti-CDV serum-neutralizing antibodies titer (1:14) and cytokine responses (upregulated TNF-α, IL-4, IL-2, and IFN-γ expression) after the third immunization. Following vaccination, the animals were challenged with a lethal CDV 5804Pe/H strain with a dose of 105.0 TCID50. Protective immune responses induced by the DNA vaccine alleviated clinical symptoms and pathological changes in CDV-infected ferrets. However, it cannot completely prevent virus replication and viremia in vivo as well as virus shedding due to the limited neutralizing antibody level, which eventually contributed to a survival rate of 75% (3/4) against CDV infection. Therefore, the improved strategies for the present DNA vaccines should be taken into consideration to develop more protective immunity, which includes increasing antigen expression or alternative delivery routes, such as gene gun injection.

Urtica fissa E. Pritz is not only an important medicinal plant for rheumatism and cough relief, but it is also an important forage plant. In this study, the complete chloroplast genome of U. fissa was assembled for the first time and reported to be 146,837 base pairs (bp) long with a typical tetragonal structure and including a large single-copy of 79,657 bp, a small single-copy of 17,712 bp, and two inverted repeats of 24,734 bp each. It harbors 115 unique genes, including 70 protein-coding genes, 38 transfer RNA genes, and 7 ribosomal RNA genes. Phylogenetic analysis showed that U. fissa is closely related to Urtica lobatifolia. This study contributes to the understanding of the origin and evolution of U. fissa, as well as its genetic relationships with other species.

Many viruses can cause infections in mink, including canine distemper virus, mink enteritis virus, and Aleutian disease virus. Current treatments are ineffective, and these infections are often fatal, causing severe economic losses. As antiviral drugs may effectively prevent and control these infections, recent research has increasingly focused on antiviral interferons. Herein, the gene encoding a mature mink interferon alpha (MiIFN-α) was synthesized according to the P. pastoris preference of codon usage and a recombinant plasmid, pPICZαA-MiIFN-α, was constructed. pPICZαA-MiIFN-α was linearized and transformed into the P. pastoris X33 strain, and zeocin-resistant transformants were selected. Protein expression was induced by methanol. SDS-PAGE and western blot analyses showed that a 25-kDa fusion protein was expressed in the culture supernatant. Antiviral activity of the expressed protein was determined using cytopathic effect inhibition (CPEI). The purified MiIFN-α significantly inhibited the cytopathic effect of vesicular stomatitis virus with a green fluorescent protein (VSV-GFP) in F81 feline kidney cells, with an antiviral activity of 6.4 × 107 IU/mL; it also significantly inhibited MEV replication in F81 cells. MiIFN-α antiviral activity against VSV-GFP was significantly reduced on treatment with pH 4 and pH 10 conditions for 24 h (p < 0.01). Serum MiIFN-α concentrations in rat were measured using enzyme-linked immune-sorbent assay; MiIFN-α concentrations in rat serum peaked at ~36 h after injection. A high dose of MiIFN-α was safe for use. There were no significant differences in body temperature, tissue changes, and lymphocyte, total white blood cell, and central granulocyte counts between the injected and control groups (p > 0.05). These findings lay a foundation for the large-scale production of recombinant MiIFNs.

BackgroundA consensus has not been made about the predictive value of blood vitamin D level in patients with coronary artery disease (CAD). This meta-analysis aimed to assess the association between blood 25-hydroxyvitamin D level and adverse outcomes in patients with CAD.MethodsTwo independent authors searched the articles indexed in PubMed and Embase databases until June 28, 2022. Cohort studies or post-hoc analysis randomized trials evaluating the value of 25-hydroxyvitamin D level in predicting cardiovascular or all-cause mortality, and major adverse cardiovascular events ([MACEs] including death, non-fatal myocardial infarction, heart failure, revascularization, stroke, etc.) were included.ResultsThe literature search identified 13 eligible studies for our analysis, including 17,892 patients with CAD. Meta-analysis showed that the pooled adjusted risk ratio (RR) was 1.60 (95% confidence intervals [CI] 1.35–1.89) for all-cause mortality, 1.48 (95% CI 1.28–1.71) for cardiovascular mortality, and 1.33 (95% CI 1.18–1.49) for MACEs. Leave-out one study sensitivity analysis suggested that the predictive values of blood 25-hydroxyvitamin D level were reliable.ConclusionsLow blood 25-hydroxyvitamin D level is possibly an independent predictor of cardiovascular or all-cause mortality and MACEs in patients with CAD. Baseline 25-hydroxyvitamin D level may provide useful information in CAD patients.

Objective We aimed to discover potential circulating genes and non-coding molecules (micro RNA [miRNA] and circular RNA [circRNA]) in CD4 + T cells in relation to seasonal allergic rhinitis (SAR). Methods Microarray data of GSE50223 were obtained from the Gene Expression Omnibus database. Differentially expressed genes (DEGs) during and outside the pollen season were analyzed using R software and by Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes pathway analyses. The protein–protein interactions, modules, miRNAs targeting DEGs, merged miRNA–DEG networks, and circRNAs targeted with miRNAs were further analyzed. Results We identified 211 DEGs during the pollen season and eight DEGs outside the season, of which only MMP12 , NR4A2 , and CD69 were differentially expressed both during and outside the pollen season. DEGs during the pollen season were enriched in the GO categories ‘neutrophil degranulation’, ‘neutrophil activation involved in immune response’, ‘neutrophil mediated immunity’, and ‘neutrophil activation’. A significant module was identified with key nodes of CDK6 and hsa-miR-29b-3p. Six significant circRNAs were also identified. Conclusions Some genes, miRNAs, and circRNAs in CD4 + T may play vital roles in SAR and may thus be potential targets for the prevention and treatment of SAR.

Microcystin-LR (MC-LR) is a toxic secondary metabolite produced by cyanobacteria that has been demonstrated to promote colorectal cancer (CRC). However, the mechanism by which MC-LR enhances CRC in the tumor microenvironment (TME) is poorly understood. To elucidate its role in TME, a co-culture system was established using CRC cells and M2 macrophages in a Transwell chamber. The study found that MC-LR promotes CRC cell migration by upregulating TGF-β1 expression and secretion in M2 macrophages and downregulating CST3 in CRC cells. Neutralizing TGF-β1 increased CST3 expression in CRC cells, while overexpressing CST3 in CRC cells suppressed TGF-β1 expression in M2 macrophages, both of which weakened MC-LR-induced cellular motility in the co-culture system. In vivo, the mice in the MC-LR/AOM/DSS group had more tumor nodules, deeper tumor invasion, and higher M2 macrophage infiltration compared to the AOM/DSS group, and the expression of TGF-β1 and CST3 in tumors was consistent with the cellular level. Overall, this study provides insights into the regulatory mechanism of MC-LR on TME, revealing that MC-LR upregulates the expression and secretion of TGF-β1 in M2 macrophages, which in turn inhibits the expression of CST3 in CRC cells to promote migration.

Abstract The Bcr/Abl plays a central role in Philadelphia chromosome‐positive (Ph+) leukemia because of the constitutively activated Abl tyrosine kinase and its downstream pathways. Currently, the clinical treatment of imatinib‐resistant patients with tyrosine kinase inhibitors is severely limited by drug resistance and adverse effects. Herein, a dual‐targeting proteolysis‐targeting chimera (PROTAC) protein drug, termed PMIBcr/Abl‐R6, is designed by engrafting an MDM2/p53 inhibition peptide sequence onto the Bcr/Abl tetramerization domain. PMIBcr/Abl‐R6, harboring a Bcr/Abl targeting sequence and an MDM2 binding sequence, acts as a PROTAC drug in Ph+ leukemia cells. Its dual‐targeting constitution suggests that PMIBcr/Abl‐R6 designs to target the tetramerization domain instead of the Abl kinase domain, therefore has the potential to overcome drug resistance mutations in the kinase domain. The efficient ability of PMIBcr/Abl‐R6 is demonstrated to simultaneously induce Bcr/Abl degradation and activate the p53 pathway. PMIBcr/Abl‐R6 has the potential to overcome drug resistance in Ph+ leukemias by multiple mechanisms.

Abstract Background Ovarian cancer (OC) is a life-threatening gynecological malignancy where dysregulation of microRNAs (miRNAs) is frequently implicated. This study focuses on the function of miR-545 on OC development and the molecules involved. Methods miR-545 expression in OC tissues and cell lines was determined, and its link to the survival of patients was analyzed. Altered expression of miR-545 was induced to determine its role in proliferation, apoptosis, migration and invasion of OC cells and the angiogenesis ability of human umbilical vein endothelial cells (HUVECs). The targeting mRNAs of miR-545 were predicted and validated through luciferase assays. Gain-of-function studies of KDM4B and PLK1 were performed to explore their involvements in OC development. In vivo experiments were conducted by inducing xenograft tumors in nude mice. Results Poor expression of miR-545 was found in OC tissues and cells compared to the normal ones and it indicated unfavorable prognosis in patients. Overexpression of miR-545 suppressed growth, migration, invasion and angiogenesis of OC cells as well as the angiogenesis ability of HUVECs. miR-545 was found to target mRNAs of KDM4B and PLK1, while KDM4B promoted the transcription of the PLK1 promoter through demethylation of H3K9me3. Either overexpression of KDM4B or PLK1 partially blocked the inhibitory effects of miR-545 mimic on OC cell growth, especially the former one. The in vitro results were reproduced in vivo. Conclusion This study evidenced that miR-545 suppresses progression of OC through mediating PLK1 expression by a direct binding and an indirect regulation involving KDM4B-mediated demethylation.

Abstract Background Aberrant activation of the Wnt/β-catenin signaling pathway is one of the most frequent abnormalities in human cancer, including colorectal cancer (CRC). Previous studies revealed pivotal functions of WNT family members in colorectal cancer, as well as their prognostic values. Nevertheless, the prognostic role and mechanisms underlying WNT7b in colorectal cancer development remains unclear. Methods In this study, WNT7b expression was measured by immunohistochemical staining of 100 cases of surgically resected human colorectal cancerous tissues as well as matched adjacent normal tissues constructed as tissue microarrays. In vitro studies, we attempted to substantiate the WNT7b expressional pattern previously found in immunohistochemistry staining. We used the colorectal cancer cell-line HCT116 and normal colorectal cell-line FHC for immunofluorescence staining and nuclear/cytoplasmic separated western blotting. We measured epithelial–mesenchymal transition (EMT) markers and migration capacity of HCT116 in the context of WNT7b knocked-down using short interfering RNA. Finally, clinical and prognostic values of WNT7b activation levels were examined. Results WNT7b was expressed in the nucleus in adjacent normal tissues. In CRC tissues, nuclear expression of WNT7b was similar; however, membrane and cytoplasmic expression was strikingly enhanced. Consistently, in vitro analysis confirmed the same expression pattern of WNT7b. Compared with FHC cells, HCT116 cells displayed higher levels of WNT7b membrane and cytoplasmic enrichment, as well as higher migration capacity with a sensitized EMT process. Either partial knockdown of WNT7b or blockade of the Wnt/β-catenin signaling pathway reversed EMT process and inhibited the migration of HCT116 cells. Finally, elevated secretion levels of WNT7b were significantly associated with lymphatic and remote metastasis and predicted worse prognosis in the CRC cohort. Conclusion In summary, we demonstrated that the activation of WNT7b autocrine probably contributes to CRC metastasis by triggering EMT process through the Wnt/β-catenin signaling pathway. High levels of WNT7b autocrine secretion predicts poor outcome in patients with CRC. This molecule is a promising candidate for clinical CRC treatments.

The aim of this study was to further broaden the heterologous expression of alginate lyase from Vibrio alginolyticus in a Bacillus subtilis expression vector. A B. subtilis WB600/pP43NMK-alg62 strain was constructed. (NH4)2SO4 precipitation and Ni-affinity chromatography were performed to purify the enzyme. We then characterized the enzyme. Its molecular weight was 57.64 kDa, and it worked optimally at 30 °C with a pH of 8.0. Ca2+ markedly enhanced the enzymatic activity of Alg62 while Cu2+ and Ni2+ inhibited its activity. Alg62 had a wide range of substrate specificity, showing high activity toward sodium alginate and polyG. Following optimization of the fermentation process, the optimal conditions for the recombinant expression of Alg62 were as follows: temperature of 37 °C, pH of 7.0, medium consisting of glycerol 15 g/L, yeast powder 25 g/L and K+ 1.5 mmol/L. At these optimal conditions, enzyme activity reached 318.21 U/mL, which was 1.54 times higher than the initial enzyme activity.

Pyruvate is a hub of various endogenous metabolic pathways, including glycolysis, TCA cycle, amino acid, and fatty acid biosynthesis. It has also been used as a precursor for pyruvate-derived compounds such as acetoin, 2,3-butanediol (2,3-BD), butanol, butyrate, and L-alanine biosynthesis. Pyruvate and derivatives are widely utilized in food, pharmaceuticals, pesticides, feed additives, and bioenergy industries. However, compounds such as pyruvate, acetoin, and butanol are often chemically synthesized from fossil feedstocks, resulting in declining fossil fuels and increasing environmental pollution. Metabolic engineering is a powerful tool for producing eco-friendly chemicals from renewable biomass resources through microbial fermentation. Here, we review and systematically summarize recent advances in the biosynthesis pathways, regulatory mechanisms, and metabolic engineering strategies for pyruvate and derivatives. Furthermore, the establishment of sustainable industrial synthesis platforms based on alternative substrates and new tools to produce these compounds is elaborated. Finally, we discuss the potential difficulties in the current metabolic engineering of pyruvate and derivatives and promising strategies for constructing efficient producers.

Helicoverpa assulta (Guenée), a moth species belonging to the Noctuidae (Lepidoptera) family, is a destructive agricultural pest that infests multiple cash crops. To assess differences in the gene expression profiles of different tissues in H. assulta, we analyzed the transcriptomes of two tissue types (midgut and hemocytes) using the Illumina Hiseq 2000 platform, on the basis of which we obtained 52076750 and 53404200 high-quality clean reads, respectively. De novo assembly yielded 46146 and 33707 unigenes from the midgut and hemocytes, respectively. After screening, we identified 23726 unigenes differentially expressed between the midgut and hemocytes. Taking the midgut as the control, we detected 7448 and 16278 unigenes that were up- and downregulated in hemocytes, respectively. Gene Ontology functional annotation divided the differentially expressed unigenes (DEUs) into three categories (biological process, cellular component, and molecular function) and 51 branches, whereas the Kyoto Encyclopedia of Genes and Genomes metabolic pathway annotation assigned the DEUs to six categories, mapping these to 258 pathways. In addition, we detected 224918 single-nucleotide polymorphic sites. Our findings based on transcriptome sequencing, data assembly, and functional gene annotation of two different tissues in H. assulta will provide a valuable reference for further excavation and study of functional genes in H. assulta.

Most studies on environmental policy and total factor productivity (TFP) growth under the heterogeneity framework tend to ignore the distance to the technical frontier, while research that investigates TFP growth based on technical distances does not tend to consider environmental policy. To fill this research gap, this study investigates the impact of environmental regulation on the total factor productivity of heterogeneous firms, based on technical distance. In addition to theoretical analysis, we apply a two-direction fixed effects model to test the impact using firm-level data selected from the CSMAR database and environmental regulation data of 287 Chinese cities between 2007 and 2015. We report two major findings from our analysis. First, environmental regulation increasingly enhances (or hinders) TFP growth, as firms get closer to (or further away from) the country-industry technology frontier, ceteris paribus. Second, grouped regression further highlights that environmental regulation affects TFP growth for heterogeneous firms. For proximal-type firms, environmental regulation promotes the growth of TFP through innovation and imitation mechanisms, while only the imitation mechanism works for middle-type firms. Neither mechanism, however, applies to distal-type firms, for whom environmental regulation hinders TFP growth. These conclusions provide a theoretical and practical basis for environmental policy, suggesting that the focus should be directed toward improving exit mechanisms for distal-type firms, creating a favorable market environment to accelerate the convergence of middle-type firms to the frontier, and encouraging proximal-type firms to innovate to catch up with or surpass the global frontier.

Objectives: Unplanned readmissions severely affect a patient’s physical and mental well-being after kidney transplantation (KT), which is also independently associated with morbidity. A retrospective study was conducted to identify the incidence, causes and risk factors for unplanned readmission after KT among Chinese patients. Methods: Patients who underwent KT were admitted to the organ transplant center of the Affiliated Hospital of University of Science and Technology of China (2017–2018). Medical records for these patients were obtained through the hospital information system (HIS). Results: In 518 patients, the incidence of unplanned readmissions within 30 days (n = 9) was 1.74%, and 90 days (n = 64) was 12.35%. The one-year unplanned readmission rate was 22.59% (n = 122). Overall, 122 patients were readmitted because of infection, renal events, metabolic disturbances, surgical complications, etc. Hemodialysis (OR = 10.462, 95% CI: 1.355–80.748), peritoneal dialysis (OR = 8.746, 95% CI: 1.074–71.238) and length of stay (OR = 1.023, 95% CI: 1.006–1.040) were independent risk factors for unplanned readmissions. Conclusion: Unplanned readmission rates increased with time after KT. Certain risk factors related to unplanned readmissions should be deeply excavated. Targeted interventions for controllable factors to alleviate the rate of unplanned readmissions should be identified.

Abstract Background Previous randomized controlled trials have compared the efficacy and safety of single chest drain (SCD) and double chest drains (DCD) in the patients undergone pulmonary lobectomy, yet the results remain inconsistent. Therefore, we aimed to conduct this present systematic review and meta-analysis to evaluate the role of SCD and DCD in the patients undergone pulmonary lobectomy. Methods PubMed, Medline, EMBASE, Cochrane library, Web of Science, China National Knowledge Infrastructure, Wanfang, Weipu, and China Biomedical Literature databases were searched up to February 28, 2020, to identify the potential RCTs on SCD and DCD in the patients undergone pulmonary lobectomy. The main outcomes including verbal pain score, the duration of drainage (days), the length of hospital stay (days), and the incidence of air leak and re-drainage were collected and analyzed. All the data were processed and analyzed with software RevMan 5.3. We calculated and analyzed the odds ratios (OR) for dichotomous outcomes and the mean difference (MD) for continuous outcomes. Results A total of 11 RCTs with 1214 patients were included, in which 589 patients received SCD treatment and 625 patients DCD treatment. The verbal pain score (MD = − 0.54, 95%CI (− 0.87, − 0.21)), the duration of drainage (MD = − 0.65, 95%CI (− 1.04, − 0.26)), and the length of hospital stay (MD = − 0.55, 95%CI (− 0.80, − 0.29)) in SCD group were significantly less than that of DCD group. There were no significant differences on the incidence of air leak (OR = 1.35, 95%CI (0.86, 2.11)) and re-drainage (OR = 0.88, 95%CI (0.41, 1.90)) among the two groups. Conclusions SCD is a safe option, and it has the advantages of less postoperative pain, shortened duration of drain, and reduced length of hospital stay when compared with DCD in the patients undergone pulmonary lobectomy.

The molecular epidemiology and biological characteristics of Escherichia coli associated with hemorrhagic pneumonia (HP) mink from five Chinese Provinces were determined. From 2017 to 2019, 85 E. coli strains were identified from 115 lung samples of mink suffering from HP. These samples were subjected to serotyping, antimicrobial susceptibility, detection of virulence genes, phylogenetic grouping, whole-genome sequencing, drug resistant gene, multilocus sequence typing (MLST) and biofilm-forming assays. E. coli strains were divided into 18 serotypes. Thirty-nine E. coli strains belonged to the O11 serotype. Eighty-five E. coli strains were classified into seven phylogenetic groups: E (45.9%, 39/85), A (27.1%, 23/85), B1 (14.1%, 12/85), B2 (3.7%, 3/85), D (3.7%, 3/85), F (2.4%, 2/85) and clade I (1.2%, 1/85). MLST showed that the main sequence types (STs) were ST457 (27/66), All E. coli strains had ≥4 virulence genes. The prevalence of virulence was 98.8% for yijp and fimC, 96.5% for iucD, 95.3% for ompA, 91.8% for cnf-Ⅰ, 89.4% for mat, 82.3% for hlyF, and 81.2% for ibeB. The prevalence of virulence genes iss, cva/cvi, aatA, ibeA, vat, hlyF, and STa was 3.5–57.6%. All E. coli strains were sensitive to sulfamethoxazole, but high resistance was shown to tetracycline (76.5%), chloramphenicol (71.8%), ciprofloxacin (63.5%) and florfenicol (52.9%), resistance to other antibiotics was 35.3–16.5%. The types and ratios of drug-resistance genes were tet(A), strA, strB, sul2, oqxA, blaTEM-1B, floR, and catA1 had the highest frequency from 34%-65%, which were consistent with our drug resistance phenotype tetracycline, florfenicol, quinolones, chloramphenicol, the bla-NDM-I and mcr-I were presented in ST457 strains. Out of 85 E. coli strains, six (7.1%) possessed a strong ability, 12 (14.1%) possessed a moderate ability, and 64 (75.3%) showed a weak ability to form biofilm. Our data will aid understanding of the epidemiological background and provide a clinical basis for HP treatment in mink caused by E. coli.

Molecular heterogeneity has great significance in the disease biology of multiple myeloma (MM). Thus, the analysis combined single-cell RNA-seq (scRNA-seq) and bulk RNA-seq data were performed to investigate the clonal evolution characteristics and to find novel prognostic targets in MM. The scRNA-seq data were analyzed by the Seurat pipeline and Monocle 2 to identify MM cell branches with different differentiation states. Marker genes in each branch were uploaded to the STRING database to construct the Protein-Protein Interaction (PPI) network, followed by the detection of hub genes by Cytoscape software. Using bulk RNA-seq data, Kaplan-Meier (K-M) survival analysis was then carried out to determine prognostic biomarkers in MM. A total of 342 marker genes in two branches with different differentiation states were identified, and the top 20 marker genes with the highest scores in the network calculated by the MCC algorithm were selected as hub genes in MM. Furthermore, K-M survival analysis revealed that higher NDUFB8, COX6C, NDUFA6, USMG5, and COX5B expression correlated closely with a worse prognosis in MM patients. Moreover, ssGSEA and Pearson analyses showed that their expression had a significant negative correlation with the proportion of Tcm (central memory cell) immune cells. Our findings identified NDUFB8, COX6C, NDUFA6, USMG5, and COX5B as novel prognostic biomarkers in MM, and also revealed the significance of genetic heterogeneity during cell differentiation in MM prognosis.

Background: The collagenase encoding gene col was cloned into a pP43NMK vector and amplified in Escherichia coli JM109 cells. The shuttle vector pP43NMK was used to sub-clone the col gene to obtain the vector pP43NMK-col for the expression of collagenase in Bacillus subtilis WB600. The enzyme was characterized and the composition of the expression medium and culture conditions were optimized. Methods: The expressed recombinant enzyme was purified by ammonium sulfate, ultrafiltration, and through a nickel column. The purified collagenase had an activity of 9405.54 U/mg. Results: The recombinant enzyme exhibited optimal activity at pH 9.0 and 50 °C. Catalytic efficiency of the recombinant collagenase was inhibited by Fe3+ and Cu2+, but stimulated by Co2+, Ca2+, Zn2+, and Mg2+. The optimal conditions for its growth were at pH 7.0 and 35 °C, using 15 g/L of fructose and 36 g/L of yeast powder and peptone mixture (2:1) at 260 rpm with 11% inoculation. The maximal extracellular activity of the recombinant collagenase reached 2746.7 U/mL after optimization of culture conditions, which was 2.4-fold higher than that before optimization. Conclusions: This study is a first attempt to recombinantly express collagenase in B. subtilis WB600 and optimize its expression conditions, its production conditions, and possible scale-up.

CRISPR/Cas9 is a valuable tool and has been extensively employed to perform gene editing in plants. However, CRISPR/Cas9 has not been successfully used in spinach, an important leafy vegetable crop. Here, we established a CRISPR/Cas9-based gene-editing system for spinach hairy roots and edited two cellulose synthase-like D (CSLD) genes (SoCSLD2 and SoCSLD3) that were involved in root-hair formation of spinach hairy roots. Four mutation types (i.e., replacement, insertion, deletion, and combined mutations) were observed, among which the replacement accounted for the vast majority (about 64.1%). Mutation rate differed largely among different targets. Seven homozygous/bi-allelic and eight heterozygous/chimeric mutants of SoCSLD2 were obtained from 15 independent transgenic hairy root lines. All of the seven homozygous/bi-allelic mutant lines displayed bulking and short root hairs, which resembled the characteristics of Arabidopsis atcsld2 mutants. The transcriptomic analysis further revealed that multiple gene expressions for cell-wall modulation and membrane trafficking were disturbed, which might result in the inhibition of root hair growth in socsld2 mutants. This indicates that SoCSLD2 was successfully knocked out in spinach root hairs using the CRISPR/Cas9-based gene editing system.

Elastin-like polypeptides (ELPs) are stimulus-responsive artificially designed proteins synthesized from the core amino acid sequence of human tropoelastin. ELPs have good biocompatibility and biodegradability and do not systemically induce adverse immune responses, making them a suitable module for drug delivery. Design strategies can equip ELPs with the ability to respond to changes in temperature and pH or the capacity to self-assemble into nanoparticles. These unique tunable biophysicochemical properties make ELPs among the most widely studied biopolymers employed in protein purification, drug delivery, tissue engineering and even in tumor therapy. As a module for drug delivery and as a carrier to target tumor cells, the combination of ELPs with therapeutic drugs, antibodies and photo-oxidation molecules has been shown to result in improved pharmacokinetic properties (prolonged half-life, drug targeting, cell penetration and controlled release) while restricting the cytotoxicity of the drug to a confined infected site. In this review, we summarize the latest developments in the application methods of ELP employed in tumor therapy, with a focus on its conjugation with peptide drugs, antibodies and photosensitizers.

The shikimate pathway is a necessary pathway for the synthesis of aromatic compounds. The intermediate products of the shikimate pathway and its branching pathway have promising properties in many fields, especially in the pharmaceutical industry. Many important compounds, such as shikimic acid, quinic acid, chlorogenic acid, gallic acid, pyrogallol, catechol and so on, can be synthesized by the shikimate pathway. Among them, shikimic acid is the key raw material for the synthesis of GS4104 (Tamiflu®), an inhibitor of neuraminidase against avian influenza virus. Quininic acid is an important intermediate for synthesis of a variety of raw chemical materials and drugs. Gallic acid and catechol receive widespread attention as pharmaceutical intermediates. It is one of the hotspots to accumulate many kinds of target products by rationally modifying the shikimate pathway and its branches in recombinant strains by means of metabolic engineering. This review considers the effects of classical metabolic engineering methods, such as central carbon metabolism (CCM) pathway modification, key enzyme gene modification, blocking the downstream pathway on the shikimate pathway, as well as several expansion pathways and metabolic engineering strategies of the shikimate pathway, and expounds the synthetic biology in recent years in the application of the shikimate pathway and the future development direction.

In this study, the fermentation broth of the recombinant Pichia pastoris strain ncy-2 was studied. After pretreatment, separation, and purification, lysozyme was optimized using biofilm and ion exchange separation. Finally, lysozyme dry enzyme powder was prepared by concentrating and vacuum drying. The removal rate of bacterial cells was 99.99% when the fermentation broth was centrifuged at low temperature. The optimum conditions were: transmembrane pressure of 0.20 MPa, pH 6.5, 96.6% yield of lysozyme, enzyme activity of 2612.1 u/mg, which was 1.78 times higher than that of the original enzyme; D152 resin was used for adsorption and elution. Process conditions were optimized: the volume ratio of resin to liquid was 15%; the adsorption time was 4 h; the concentration of NaCl was 1.0 mol/L; the recovery rate of lysozyme activity was 95.67%; the enzyme activity was 3879.6 u/mL; and the purification multiple was 0.5, 3.1 times of the original enzyme activity. The enzyme activity of lysozyme dry enzyme powder was 12,573.6 u/mg, which had an inhibitory effect on microsphere lysozyme. Its enzymatic properties were almost the same as those of natural lysozyme, which demonstrated good application prospects and production potential.

In the process of acquiring mutants mediated by CRISPR/Cas9, plantlets are often regenerated from both mutated and non-mutated cells in a random manner, which increase the odds of chimeric mutated plant. In general, it’s necessary to infect more explants or grow to next generation for the need of generating more biallelic or homozygous mutants. In present study, an efficient way of obtaining biallelic or homozygous mutated lines via fast-growing hairy root system without increasing numbers of infected explants or prolonging sexual propagation generation is reported. The fast growing lateral branches of hair roots are originated deep within the parental root from a small number of founder cells at the periphery, and therefore were employed as a library that classify different editing types in different lateral branches in which the homozygous or biallelic lines were screened. Here, MtPDS was employed in a proof-of-concept experiment to evaluate the efficiency of genome editing with our hairy root system. Homozygous/biallelic mutations were found only 1 of the 20 lines in the 1st generation hairy roots, and 8 lines randomly selected were cultured to obtain their branch roots, homozygous/biallelic mutations were found in 6 of the 8 lines in their branch roots. We also tested the method with MtCOMT gene and got the same result. All of the seedlings regenerated from the homozygous/biallelic hairy root mutation lines of MtPDS displayed albino phenotypes. The entire process from vector design to the recovery of plantlets with homozygous/biallelic mutations took approximately 4.5–6.5 months. The whole process could bring inspiration for efficiently generating homozygous/biallelic mutants through CRISPR/Cas9 system from the hairy root or root system of a chimeric mutated transformants, especially for the rare and endangered plants whose explants sources are very limited or the plants that lack of tissue culture and rapid propagation system.

We presented a patient with CML who progressed to unclassical blast phase after Tyrosine Kinase Inhibitors (TKIs) therapy. The patient presented with 2 populations of blasts: one with no cytoplasmic granules and was CD117 weak+/tryptase-/CD34- (typical myeloblasts), and another with metochromatic granules in the cytoplasm and was CD117 strong+/tryptase+/CD25+/CD34 subset+ (myelomastocytic blasts). Almost all the cells were positive for BCR/ABL1 fusion and no KT V816F mutation was detected. The patient was misdiagnosed as having blast phase CML with coexisting mast cell leukemia at an outside institute. Three similar cases and previously described myelomastocytic leukemia are reviewed and discussed.