[Mechanism of Qilongtian Capsules in treatment of acute lung injury based on network pharmacology prediction and experimental validation].

This study aimed to explore the mechanism of Qilongtian Capsules in treating acute lung injury(ALI) based on network pharmacology prediction and in vitro experimental validation. Firstly, UPLC-Q-TOF-MS/MS was used to analyze the main chemical components of Qilongtian Capsules, and related databases were used to obtain its action targets and ALI disease targets. STRING database was used to build a protein-protein interaction(PPI) network. Metascape database was used to conduct enrichment analysis of Gene Ontology(GO) and Kyoto Encyclopedia of Genes and Genomes(KEGG). AutoDock software was used to perform molecular docking verification on the main active components and key targets. Then, the RAW264.7 cells were stimulated with lipopolysaccharide(LPS) for in vitro experiments. Cell viability was measured by MTT and ROS level was measured by DCFH-DA. NO content was measured by Griess assay, and IL-1ß, IL-6, and TNF-α mRNA expression was detected by RT-PCR. The predicted targets were preliminarily verified by investigating the effect of Qilongtian Capsules on downstream cytokines. Eighty-four compounds were identified by UPLC-Q-TOF-MS/MS. Through database retrieval, 44 active components with 589 target genes were screened out. There were 560 ALI disease targets, and 65 intersection targets. PPI network topology analysis revealed 10 core targets related to ALI, including STAT3, JUN, VEGFA, CASP3, and MMP9. KEGG enrichment analysis showed that Qilongtian Capsules mainly exerted an anti-ALI effect by regulating cancer pathway, AGE-RAGE, MAPK, and JAK-STAT signaling pathways. The results of molecular docking showed that the main active components in Qilongtian Capsules, including crenulatin, ginsenoside F_1, ginsenoside Rb_1, ginsenoside Rd, ginsenoside Rg_1, ginsenoside Rg_3, notoginsenoside Fe, notoginsenoside G, notoginsenoside R_1, notoginsenoside R_2, and notoginsenoside R_3, had good binding affinities with the corresponding protein targets STAT3, JUN, VEGFA, CASP3, and MMP9. Cellular experiments showed that Qilongtian Capsules at 0.1, 0.25, and 0.5 mg·mL~(-1) reduced the release of NO, while Qilongtian Capsules at 0.25 and 0.5 mg·mL~(-1) reduced ROS production, down-regulated mRNA expression of IL-1ß, IL-6, TNF-α, and inhibited the inflammatory cascade. In summary, Qilongtian Capsules may exert therapeutic effects on ALI through multiple components and targets.

Reproductive outcomes of ectopic pregnancy with conservative and surgical treatment: A systematic review and meta-analysis.

BACKGROUND: Ectopic pregnancy (EP), one of the most common gynecological emergencies, is the major cause of maternal death in the first trimester and increases the incidence of infertility and repeat ectopic pregnancy (REP). The aim of this study was to compare the effects of different treatment methods for tubal EP on natural pregnancy outcomes. METHODS: We systematically searched PubMed, Embase, Cochrane Library, Web of Science, and Clinical Trials for observational studies on EP (published until October 30,2022 in English) comparing methotrexate (MTX) versus surgery, MTX versus salpingostomy, MTX versus salpingectomy, salpingostomy versus salpingectomy, and MTX versus expectant treatment. Our main endpoints included subsequent natural intrauterine pregnancy (IUP) and REP. We assessed the pooled data using Review Manager software (version 5.3) with a random effects model. RESULTS: Of 1274 identified articles, 20 were eligible and 3530 participants were included in our analysis. There was a significant difference in the odds of subsequent IUP in tubal EP patients who underwent MTX compared with those who were treated with surgery [odds ratios (OR) = 1.52, 95% confidence interval (CI):1.20-1.92]. No significant difference was found in the odds of REP between the 2 groups (OR = 1.12, 95% confidence interval [CI]: 0.84-1.51). There was no significant difference in the odds of subsequent IUP and REP in patients after MTX compared to those after salpingostomy (OR = 1.04,95% CI: 0.79-1.38; OR = 1.10, 95% CI: 0.64-1.90). There was a significant difference in the odds of subsequent IUP in patients after MTX compared with those after salpingectomy (OR = 2.11, 95% CI: 1.52-2.93). No significant difference was found in the odds of REP between the 2 groups (OR = 0.98, 95% CI: 0.57-1.71). There was a significant difference in the odds of subsequent IUP between patients who underwent salpingostomy and those who underwent salpingectomy (OR = 1.61, 95% CI: 1.29-2.01). No significant difference was found in the odds of REP between the 2 groups (OR = 1.21, 95% CI: 0.62-2.37). There was no significant difference in the odds of subsequent IUP and REP in patients after MTX compared with those after expectant treatment (OR = 1.25, 95% CI: 0.64-2.45; OR = 0.69, 95% CI: 0.09-5.55). CONCLUSION: For hemodynamically stable tubal EP patients, MTX has advantages over surgery, particularly salpingectomy, in improving natural pregnancy outcomes. However, MTX is not inferior to salpingostomy and expectant treatment.

[The Risk and Survival Analysis of Multiple Malignancies in Hematologic Malignancy Patients: A Single Chinese Center Retrospective Study, 2009 through 2017].

OBJECTIVE: To explore the risk and location of multiple malignancies in patients with hematologic malignancies who were followed up for 9 years in Jiangsu Province Hospital and to evaluate the impact of the second primary malignancy on survival of patients. METHODS: The incidence and survival of multiple malignancies in 7 921 patients with hematologic malignancies from 2009 to 2017 were analyzed retrospectively. RESULTS: A total of 180 (2.3%, 180/7 921) patients developed second malignancy, of whom 58 patients were diagnosed with hematologic malignancies as the first primary malignancy, and 98 patients developed hematologic malignancies as second primary malignancy, and the other 24 cases were diagnosed with the second malignancy within 6 months after the first primary malignancy was diagnosed, which was difined as multiple malignancies occurring simultaneously. In 180 patients, 18 cases developed two hematologic malignancies successively, and 11 patients developed more than 3 primary cancers (among them, 2 female patients were diagnosed with 4 primary cancers). Patients with lymphoma and multiple myeloma (MM) as the second primary malignancy had poorer survival than patients with lymphoma and MM as the first primary malignancy. Patients with chronic myeloid leukemia as the second primary malignancy were also associated with inferior overall survival. CONCLUSION: In this study, 2.3% of hematologic malignancy patients had multiple mali-gnancies, lymphoma and MM as the second primary malignancy had poor survival.

Which patients with hydrosalpinx will benefit more from reproductive surgery to improve natural pregnancy outcomes?: A systematic review and meta-analysis.

BACKGROUND: The most common tubal disease leading to infertility occurs in the distal region, manifesting as hydrosalpinx. Tubal surgery is an effective alternative treatment. However, subpopulations that benefit the most from tubal repair surgery remain unclear. The objective of this study was to investigate the natural pregnancy outcomes of patients with hydrosalpinx after reproductive surgery and those with different grades of hydrosalpinx. METHODS: We searched the major online databases (PubMed, Embase, Cochrane Library, Web of Science, and Clinical Trials) to collect observational studies on patients with hydrosalpinx who underwent surgeries to preserve natural fertility from January 2000 to August 2022. The outcome indicators were natural intrauterine pregnancy (IUP) and ectopic pregnancy (EP) rates. Studies on patients with hydrosalpinx who underwent laparoscopic surgeries and those who intended to be conceived naturally were included. Studies on patients with non-hydrosalpinx diseases, those who underwent open surgery, and those who intended to undergo assisted conception were excluded. The Newcastle-Ottawa scale for observational studies was used for quality evaluation. Meta-analysis of a single rate was performed using RevMan5.3 software. RESULTS: A total of 10 articles were included in this study, including 1317 patients with hydrosalpinx. Seven studies were retrospective and 3 were prospective. It was found that after surgery for preserving natural fertility function, the IUP and EP rates of patients with hydrosalpinx were 27% (95% confidence interval [CI]: 22-32%) and 4.8% (95% CI: 2.91-8.26%), respectively. In addition, the IUP and EP rates in patients with mild (n = 254), moderate (n = 252), and severe (n = 473) hydrosalpinx were 50.5% (95% CI: 38.65-61.97%), 32.9% (95% CI: 21.88-46.24%), 10.7% (95% CI: 4.76-21.88%), and 7.4% (95% CI: 2.91-19.35%), 9.09% (95% CI: 6.54-13.79%), 8.3%, 8.26% (95% CI: 3.85-18.03%), respectively. CONCLUSION: Patients with mild to moderate hydrosalpinx will benefit more from reproductive surgery to improve natural pregnancy outcomes. However, the small sample size in our study needs to be further expanded, and the grouping needs to be more refined, such as grouping based on age. This may provide more guidance in clinical practice.

Synthesis and Characterization of Donor-Acceptor Iron Porphyrin Carbenes and Their Reactivities in N-H Insertion and Related Three-Component Reaction.

Iron porphyrin carbenes (IPCs) have been extensively recognized as the reactive intermediates in various iron porphyrin-catalyzed carbene transfer reactions. While donor-acceptor diazo compounds have been frequently used for such transformations, the structures and reactivities of donor-acceptor IPCs are less explored. To date, no crystal structures of donor-acceptor IPC complexes have been reported, and therefore, the involvement of IPC intermediacy for such transformations lacks direct evidence. Here we report the synthesis and NMR characterization of several donor-acceptor IPC complexes from iron porphyrin and corresponding donor-acceptor diazo compounds. The X-ray crystal structure of an IPC complex derived from a morpholine-substituted diazo amide was obtained. The carbene transfer reactivities of those IPCs were tested by the N-H insertion reactions with aniline or morpholine as well as the three-component reaction with aniline and γ,δ-unsaturated α-keto ester based on electrophilic trapping of an ammonium ylide intermediate. Based on these results, IPCs were identified as the real intermediates for iron porphyrin-catalyzed carbene transfer reactions from donor-acceptor diazo compounds.

[A Retrospective Analysis of TKI Discontinuation in Patients with CML].

OBJECTIVE: To investigate the clinical characteristics of the patients with chronic myeloid leukemia (CML) discontinued tyrosine kinase inhibitors (TKI) therapy and the outcome of the patients. METHODS: 35 cases of CML patients experienced initiative discontinuation of TKI therapy in our hospital from June 1st 2015 to December 31th 2019 were retrospectively analyzed. The TFR of the patients and the factors affecting it were analyzed. RESULTS: The median duration of TKI administration was 72 (range 35-173) months in the 35 patients. Among these patients, 8 had experienced TKI dose reduction or suspension. All the enrolled patients have achieved at least MMR. The median time for these patients achieving MMR was 15 (range 3-75) months after administration of TKI, and for MMR maintenance before TKI suspension was 55 (range 13-164) months. After TKI withdrawal the median follow up time was 20.3 (range 3-57.9) months, 22 out of 35 patients kept TFR, among them, 2 （5.71%） patients restarted TKI after 12 month suspension, and maintained MMR during suspension. 13 （37.1%）patients lost MMR, among them, 9 patients restarted TKI treatment, and 5 of them achieved MR4.0 after the median duration of 3(2-5) month. No patients were found to have disease progression. The estimated TFR rate was 57.8% and 51.8% at 12 and 24 months after discontinuation, respectively. Other clinical characteristic related to relapse were also analyzed, including the cumulative TKI administration duration, cumulative MMR duration, time to achieve MMR, median age at diagnosis, risk stratification by Sokal score, TKI dose reduction and discontinuation history, and second-generation TKI administration before stopping TKI, however, no statistical difference was found. CONCLUSION: TKI discontinuation is practical for CML patients in our center.

[Application of Various Methods to Extract Microplastic from Typical Soils in China].

Microplastics have been found in many environmental systems, such as oceans, terrestrial soils, sediments, and bodies of freshwater. Microplastic pollution in soils has received extensive international attention; however, there is currently no unified standard extraction method. To identify appropriate extraction and component identification methods for microplastics in typical soils in China, samples were identified and their polymer component properties examined using ATR-FTIR(Thermo Nicolet IS 20). The effects of three treatments of water+oil(T1), a saturated NaCl solution+oil(T2), and a saturated NaCl solution(T3) on the detection of four types of microplastics[polyethylene terephthalate(PET), polypropylene(PP), polystyrene(PS), and polyethylene(PE)]in four typical soils(loess, black soil, red soil, and purple soil) were examined. The results showed that:① The extraction rates of the T3 treatment decreased with microplastic density. For PP, PE, and PS, the extraction rates exceeded 86.67%, and the extraction rates of PET were 0%-13.30%. ② The extraction rates of the four kinds of microplastics were 86.67%-100.00% in loess, black soil, and purple soil using the T1 and T2 solutions; however, the residual oil on the surface of microplastics influenced the identification using FTIR. Furthermore, the cleaning effect of PE and PS was poor after wiping with anhydrous ethanol. ③ In red soil, the extraction rates of PET were 56.60% and 50.00% in T1 and T2, respectively; in the T3 treatment, the extraction rates of PET, PE, and PP were 3.33%, 10.00%, and 56.67%, respectively. These results show that the extraction rates of four types microplastics are highest for loess, black soil, and purple soil using T1 and T2, although these two methods need to be combined with anhydrous ethanol cleaning. T3 dispose making the leak of microplastics PET with the density higher than 1 g·cm-3. Methods to extract microplastics from red soils require further research. Overall, this study can inform the investigation and treatment of microplastic pollution in typical regional soils in China.

Fertility-preserving treatment in patients with early-stage endometrial cancer: A protocol for systematic review and meta-analysis.

BACKGROUND: Endometrial cancer (EC) is the second most common malignancy of the female reproductive system worldwide, and the standard treatment for early-stage EC potentially leads to permanent infertility. The objective of this study was to investigate the efficacies of different methods on fertility preservation in patients with early-stage EC. METHODS: We searched the major online databases (PubMed, Embase, The Cochrane Library, and Web of Science) to collect the research literature on fertility preservation therapy in patients with early-stage well-differentiated EC aged ≤ 40 years from January 1999 to October 2019. The inclusion was performed using the R software (version R3.5.3) meta-analysis of a single rate. The efficacy of the following three fertility preservation treatments was evaluated from four aspects, the complete remission rate (CRR), recurrence rate (ReR), pregnancy rate (PregR), and live birth rate (LBR): a) taking oral progestin only therapy, b) hysteroscopic resection combined with progestin/levonorgestrel-releasing intrauterine system (LNG-IUS)/GnRH-a, c) LNG-IUS or combined with progestin/GnRH-a. RESULTS: A total of 23 articles were included in this study, including 446 patients with early-stage EC. In the group that took oral progestin only (n = 279), CRR, ReR, PregR, and LBR were 82% (95% confidence interval [CI], 74%-92%, P = .01), 38% (95% CI, 31%-45%, P = .35), 70% (95% CI, 62%-79%, P = .68), and 63% (95% CI, 55%-73%, P = .55), respectively. Hysteroscopic resection combined with progestin/LNG-IUS/GnRH-a therapy group (n = 96) achieved a CRR, ReR, PregR, and LBR of 95% (95% CI, 90%-100%, P = .42), 16% (95% CI, 6%-39%, P = .03), 84% (95% CI, 73%-96%, P = .39), and 72% (95% CI, 59%-87%, P = .28), respectively. LNG-IUS or combined with progestin/GnRH-a therapy group (n = 91) achieved a CRR, ReR, PregR, and LBR of 69% (95% CI, 54%-89%, P < .01), 30% (95% CI, 19%-49%, P = .36), 48% (95% CI, 18%-100%, P < .01), and 36% (95% CI, 10%-100%, P < .01), respectively. CONCLUSION: It is safe and effective for young patients with early-stage EC to receive oral progestin, hysteroscopic resection combined with progestin/LNG-IUS/GnRH-a, LNG-IUS, or progestin/GnRH-a. INPLASY REGISTRATION NUMBER: DOI 10.37766/inplasy2020.12.0137.

TANK-binding kinase 1 mediates osteoclast differentiation by regulating NF-κB, MAPK and Akt signaling pathways.

TANK-binding kinase 1 (TBK1) belongs to the noncanonical IκB kinase (IKK) family. The ubiquitously expressed protein is well known to play a pivotal role in innate immune response and inflammation. Although excessive inflammatory activities have been shown to affect osteoclast (OC) differentiation and function, direct relevance of TBK1 in bone turnover is not known. In this work, we specifically altered the TBK1 protein level by knocking down or overexpressing it without affecting its homologous protein IKKε expression, and demonstrated the effect of TBK1 on OC differentiation in bone marrow macrophages (BMMs) and RAW264.7 cells upon induction by receptor activator of nuclear factor-κB (NF-κB) ligand (RANKL). TBK1 knockdown was found to markedly inhibit the OC differentiation and function, while TBK1 overexpression enhanced OC formation. Downregulation of TBK1 greatly suppressed RANKL-induced gene expression of Mmp9, Atp6v0d2, Acp5, Ctsk andNfatc1 involved in the regulation of OC formation and function in both BMM and RAW264.7 cells. Mechanistic studies indicated that TBK1 affected the NF-κB signaling pathway as well as mitogen-activated protein kinases (MAPKs) and protein kinase B (Akt) activation during OC differentiation. Moreover, the protein level of TNF receptor-associated factor 6 (TRAF6) was increased, and the interaction of TRAF6 with TBK1 was potentiated, by RANKL. Collectively, we provide direct evidence showing that TBK1 effectively mediates OC differentiation and function by regulating NF-κB, MAPKs and Akt signals. A TBK1-targeted therapeutic strategy may be useful for the treatment of bone-related disorders.

[Clinical application of midpiece facial nerve dissection in regional parotidectomy].

OBJECTIVE: To propose and evaluate the clinical effect of midpiece facial nerve dissection through transparotid approach in regional parotidectomy. METHODS: A total of 136 patients with benign parotid tumors were categorized into three groups according to the way of facial nerve dissection: anterograde dissection from main trunk (anterograde, n=70), retrograde dissection from distal branches (retrograde, n=34), and midpiece dissection through transparotid approach (middle dissection, n=32). Surgery duration, facial nerve injury, salivary fistula, earlobe sensation, Frey's syndrome, and aesthetic evaluation were compared. RESULTS: The surgery duration in the middle dissection group was significantly shorter than that in the other two groups. The proportion of salivary fistula was higher in the anterograde group (9 cases, 12.9%; P<0.05) compared with that in the other groups. Postoperative facial nerve injury was similar between the middle dissection (1 case, 3.1%) and anterograde groups (3 cases, 4.3%) with lower injury rate compared with the retrograde group (7 cases, 20.6%). The anterograde group had more cases of hypoesthesia of the earlobe (12 cases, 17.1%; P<0.05) than the other two groups. Aesthetic score was higher in the anterograde and middle dissection groups compared with that in the retrograde group (P<0.05). CONCLUSIONS: Midpiece facial nerve dissection is technically feasible and clinically viable in regional parotidectomy.

Robust and nanoparticle-free superhydrophobic cotton fabric fabricated from all biological resources for oil/water separation.

Traditional superhydrophobic cotton fabrics (SCFs) for oil/water separation were usually fabricated by surface coating with inorganic nanoparticles combined with nonrenewable and nonbiodegradable or even toxic fossil-based chemicals, which would lead to secondary environmental pollution after their lifetime. In this study, we report robust, nanoparticle-free, fluorine-free SFC, which was prepared by acid etching followed by surface coating with epoxidized soybean oil resin (CESO) and subsequent modification with stearic acid (STA). No toxic compound and no nanoparticle were included within the SCF and all the raw materials including cotton fabric, CESO and STA are biodegradable and derived from biological resources. The SCF showed excellent mechanical stability and chemical/environmental resistances. The superhydrophobicity of the SFC survived from mechanical abrasion, tape peeling, ultrasonication, solvent erosion and low/high temperature exposure. The SCF also exhibited good acid/alkali resistance with contact angle over 150° toward different pH water droplets. Moreover, the SCF could efficiently separate oil/water mixtures with efficiency above 97.9% and the superhydrophobicity remained after reusing for at least 10 times. The fully biological-derived SCF with excellent mechanical and chemical resistances exhibit great potential for separation of oil/water mixtures.

T63 inhibits osteoclast differentiation through regulating MAPKs and Akt signaling pathways.

Inhibition of excessive osteoclast differentiation and activity is a valid approach for the treatment of osteoporosis. T63 is a small-molecule compound identified from a high throughput screening based on RUNX2 transcriptional activity, and has been reported to stimulate osteoblast formation. However, whether the compound has any effect on osteoclast differentiation remains unknown. Here, we examined the in vitro effect of T63 on osteoclastogenesis. T63 was found to inhibit the number of TRAP-positive cells in an osteoblast-osteoclast co-culture system, and inhibited Rankl expression in the preosteoblast MC3T3-E1 cells. The compound also directly suppressed RANKL-induced osteoclast differentiation in both dose- and time-dependent manner, as evidenced by the decrease of TRAP activity, F-actin formation and osteoclastogenesis-related genes expression in RAW264.7 cells. Moreover, pretreatment with T63 markedly decreased the activation of mitogen-activated protein kinases and Akt, both of which are positively involved in the regulation of osteoclastogenesis. Collectively, our findings suggest T63 has a protective effect against bone loss by inhibiting bone resorption. Its regulatory effect on bone metabolism makes the compound a more promising candidate for the potential application in the treatment of osteoporosis.

[Analysis of Hematopoietic Reconstitution in Auto-Peripheral Blood Hematopoietic Stem Cell Transplantation].

OBJECTIVE: To explore the factors which may have influences on hematopoietic reconstitution of the auto-peripheral blood stem cell transplantation(auto-PBHSCT). METHODS: The successful rate, the time of hematopoietic reconstitution and implantation status at 28 days after transplantation of 177 patients received auto-PBSCT were retropectively analyzed, in order to explore the factors which may have influences on hematopoietic reconstitution. RESULTS: The median time of neutrophil recovery was 12 days (8-21 days), implantation rate was 98.9%, all patients' neutrophil were recovered in 28 days. The median time of platelet recovery was 17 days (7-420 days), implantation rate was 95.5%, the cumulative incidence of platelet recovery at day 28 was 80.8%. Univariate analysis showed that the CD34+ cell number and the use of TPO had effect on neutrophils recovery time; the disease kinds, conditioning regimen and the infused CD34+ cell number had influence on platelets recovery time. Multivariate analysis showed that the CD34+ cell number was the independent influencing factor of neutrophils reconstitution time; the disease kinds, the CD34+ cell number were the independent influencing factors of platelet reconstitution time. Disease kinds and the CD34+ cell number were the independent influencing factors of hematopoietic reconstitution status of 28 days after transplantation. CONCLUSION: In auto-PBHSCT patients, disease kinds, conditioning regimen, the infused CD34+ cell number and the use of TPO have been confirmed to be independent influencing factors on hematopoietic reconstitution.

Aesculin modulates bone metabolism by suppressing receptor activator of NF-κB ligand (RANKL)-induced osteoclastogenesis and transduction signals.

Aesculin (AES), a coumarin compound derived from Aesculus hippocasanum L, is reported to exert protective role against inflammatory diseases, gastric disease and cancer. However, direct effect of AES in bone metabolism is deficient. In this study, we examined the effects of AES on osteoclast (OC) differentiation in receptor activator of NF-κB ligand (RANKL)-induced RAW264.7 cells. AES inhibits the OC differentiation in both dose- and time-dependent manner within non-toxic concentrations, as analyzed by Tartrate Resistant Acid Phosphatase (TRAP) staining. The actin ring formation manifesting OC function is also decreased by AES. Moreover, expressions of osteoclastogenesis related genes Trap, Atp6v0d2, Cathepsin K and Mmp-9 are decreased upon AES treatment. Mechanistically, AES attenuates the activation of MAPKs and NF-κB activity upon RANKL induction, thus leading to the reduction of Nfatc1 mRNA expression. Moreover, AES inhibits Rank expression, and RANK overexpression markedly decreases AES's effect on OC differentiation and NF-κB activity. Consistently, AES protects against bone mass loss in the ovariectomized and dexamethasone treated rat osteoporosis model. Taken together, our data demonstrate that AES can modulate bone metabolism by suppressing osteoclastogenesis and related transduction signals. AES therefore could be a promising agent for the treatment of osteoporosis.

EZH2 mediates lidamycin-induced cellular senescence through regulating p21 expression in human colon cancer cells.

Lidamycin (LDM) is a novel member of the enediyne antibiotics identified in China with potent antitumor activity. However, it remains unclear whether LDM has potential molecular targets that may affect its antitumor activity. Enhancer of zeste homolog 2 (EZH2) functions as a histone lysine methyltransferase and mediates trimethylation on histone 3 lysine 27 (H3K27me3). High EZH2 level is found to be positively correlated with the aggressiveness, metastasis and poor prognosis of cancer. Here, we aim to study the role of EZH2 in LDM-induced senescence, as well as in the cytotoxicity of LDM in human colon cancer cells. LDM is found to be relatively more potent in inhibiting the colon cancer cells harboring high EZH2 level and induces irreversible cellular senescence at IC50 dose range, as evidenced by senescence-associated ß-galactosidase staining, cell cycle arrest and molecular changes of senescence regulators including p21 in HCT116 and SW620 cells. More importantly, LDM is found to markedly inhibit EZH2 expression at both protein and mRNA levels upon the induction of p21 and cellular senescence. LDM also selectively inhibits EZH2 expression as compared with other histone lysine methyltransferases. Knockdown of p21 with siRNAs abolishes LDM-induced senescence, whereas EZH2 knockdown markedly increases p21 expression and causes senescent phenotype. Enrichment of both EZH2 and H3K27me3 levels in the p21 promoter region is reduced by LDM. Moreover, EZH2 overexpression reduces cellular senescence, p21 expression and DNA damage response upon LDM exposure. LDM also demonstrates potent antitumor efficacy in xenografted animal models. Collectively, our work provides first demonstration that EZH2 may mediate, at least partially, the senescence-inducing effects of LDM by regulating p21 expression and DNA damage effect. Thus, EZH2 may serve as a potential target and biomarker to indicate the clinical efficacy of the potent enediyne antitumor drug.

[Application of CRISPR/Cas9 Gene Editing Technique to Establish S1PR5 Gene Knockout Mice].

OBJECTIVE: To establish the S1PR5 gene knockout mouse model by using CRISPR/Cas9 gene editing technique so as to provide the tool for studying the regulating role of sphingosine-1-phosphate receptor 5 (S1PR5) in allogeneic hematopoietic stem cell transplantation. METHODS: Single guide RNA (sgRNA) plasmids against the exon 3 of S1PR5 were designed and constructed. Then the sgRNA and hCas9 were transcribed by T7 RNA polymerase in vitro. Cas9 mRNA and sgRNA were mixed and microinjected into fertilized eggs of C57BL/6 mice. T7E1 digestion and gene sequencing were used to detect the mutations of S1PR5. Quantitative PCR (qPCR) and Western blot were used to detect the expression of S1PR5. RESULTS: Finally 2 kinds of F2 generation of homozygous S1PR5 deficent mice (S1PR5-170/-170 mice and S1PR5-215/-215 mice) were gained, and in these 2 model mice the S1PR5 did not express at mRNA and protein levels. CONCLUSION: A mouse model with S1PR5 dificiency has been successfully established, which shall lay a foundation for future investigation of S1PR5.

[Influence of S1PR5 Defect on the Lymphocyte Distribution in Mice].

BACKGROUND: The sphingosine 1-phosphate (S1P) receptors (S1PRs) are a group of G protein-coupled receptors expressed on the surface of lymphocytes. The interaction between S1P and S1PRs plays a significant role in the migration and distribution of lymphocytes. OBJECTIVE: To investigate the influence of S1PR5 defect on the lymphocytes distribution in mice. METHODS: The distribution of different subsets of lymphocyte in the mice with S1PR5 defect was examined by flow cytometry. RESULTS: Compared with wild type mice, the number of NK cells in the peripheral blood (PB) and spleen (SP) from the mice with S1PR5 defect decreased very significantly 〔PB: 6.4±0.45% vs 2.2±0.47(P<0.01,n=3)；SP: 3.0±0.91% vs 0.68±0.14%(P<0.05,n=3)〕. However, the NK cell number in the bone marrow (BM) and lymphonodes (LN) of the mice with S1PR5 defect increased very significantly 〔BM: 0.97±0.20 % vs 2.6±0.35% (P<0.01, n=3); LN: 0.35±0.16% vs 1.7±0.15% (P<0.01, n=3)〕. The percentages of CD3(+) lymphocyte in peripheral blood, spleen and lymph node were not statistically significantly different between these 2 types of mice 〔PB: 17.3±7.9% vs 17.0±4.6% (P>0.05, n=3); SP: 33.0±6.0% vs 27.4±1.8% (P>0.05, n=3); LN: 42.3±10.7% vs 51.2±2.7% (P>0.05, n=3)〕. CONCLUSION: S1PR5 defect can significantly influence the NK cell distribution.

[Therapeutic Efficacy Analysis of Allogeneic Peripheral Blood Hematopoietic Stem Cell Transplantation for 14 Adult Patients with T Lymphoblastic Lymphoma].

OBJECTIVE: To investigate the therapeutic efficacy of allogeneic peripheral blood hematopoietic stem cell transpdantation (allo-HSCT) for T lymphoblastic lymphoma (T-LBL). METHODS: The clinical data of 14 adult patients with T-LBL treated with allo-HSCT were collected, the hematopoietic reconstruction, survival and relapse, as well as overall survival (OS) rate, event-free survival (EFS) rate of 1, 3 and 5 years were analysed retrospectively. RESULTS: All the patients were engrafted with neutrophil successfully, the median time of absolute neutrophil count >0.5 × 10(9)/L was 13 (10-19) d; 13 patients were engrafted with platelets successfully, the median time of Plt count >20 × 10(9)/L was 17 (12-62) days. The acute GVHD occurred in 6 patients, but among them only 1 case with 3 grade of aGVHD; out of 14 patients, 5 developed chronic GVHD. The transplant-related mortality at 100 days was 7.1% (1/14), mainly from coronary heart disease and pulmonary infection. The median follow-up time was 26.5 months, the estimated 1, 3 and 5 year OS rate was 85.7%, 47.6% and 38.1%, respectively, and estimated 1, 3 year EFS rate was 85.7%, 34.4% and 34.1%, respectively. The relapse rate was 42.8% (6/14) and the median relapse time was 22.5% months after transplantation. Up to now, 7 patients still survive, 1 patient out of them have survived for 103 months. CONCLUSION: The allo-HSCT is a safe and effective method for treatment of T-LBL.

[Clinic Outcome of Allogeneic Stem Cell Transplantation in 45 Patients with MDS].

OBJECTIVE: To investigate the clinical features and prognosis of patients with myelodysplastic syndrome (MDS) who received allogeneic hematopoietic stem cell transplantation (allo-HSCT). METHODS: A total of 45 patients with MDS and transformed acute myeloid leukemia (tAML) who received allo-HSCT between January 2009 and December 2014 were enrolled in this study. The effects of different conditioning regimens, donor and chemotherapy before transplantation on the clinical outcome were analyzed retrospectively. RESULTS: The median follow-up time for these patients was 54.6 months (ranged from 1 to 72.1 months), the 4-year cumulative overall survival (OS) and disease-free survival (DFS) rates were 77.1% and 62.1%, respectively. In myeloblative conditioning group and reduced intensity conditioning group, the 3-year cumulative OS rate was 69% and 68.6% (HR = 1.0, P = 0.984), respectively, the 3-year cumulative relapse rate was 17.6% and 33.3% (HR = 3.389, P = 0.162). The 100-day cumulative rate of aGVHD (38.6%) in HLA-identical nonsibling group was similar to HLA identical sibling group (37%) (HR = 1.089, P = 0.885); meanwhile the similar 3-year commulative OS rate was achieved in the 2 groups (72.7% and 70%) (HR = 0.952, P = 0.942). Among 26 patients with RAEB-2 and t-AML, the 2-year cumulative OS were 66.7% and 58.3% (HR = 1.265, P = 0.750) and 2-year cumulative relapse rates were 20.0% and 12.5% (HR = 0.417, P = 0.477) in non-chemotherapy and CR post-chemotherapy subgroups. The 1-year cumulative OS rate was 53.5% and 84.8% in the group with or without aGVHD. The patients with aGVHD had higher transplantation related mortality (TRM) compared with patients without aGVHD (HR = 15.0, P =0.011). CONCLUSION: The reduced intensity conditioning doesn't reduce OS rate in patients with MDS, and elderly patients can benefit from it. The OS rate is similar between HLA-identical sibling and HLA-identical nonsibling allo-HSCT. The chemotherapy before transplantation cannot prolong the survival of MDS patients.

Elevated absolute monocyte count predicts unfavorable outcomes in patients with angioimmunoblastic T-cell lymphoma.

This study was aimed at investigating the prognostic significance of the absolute monocyte count (AMC) in peripheral blood in patients with newly diagnosed angioimmunoblastic T cell lymphoma (AITL). AMC was performed in 73 therapy-naive patients with AITL in 2 institutions during 2008-2015, and higher AMC was observed in those with extranodal sites >1, bone marrow involvement, high lactate dehydrogenase level, the EBV infection, no response to treatment and high IPI, PIT, PIAI score group. The best AMC cut-off level at diagnosis was 0.8 × 10(9)/L and the 3-year overall survival (OS) was 64% for patients with low AMC group (≤ 0.8 × 10(9)/L) compared to 10% in high AMC group (>0.8 × 10(9)/L) (P<0.001). Multivariate analysis showed that elevated AMC remained an adverse prognostic parameter. Our results suggest that AMC is an independent prognostic parameter for OS in patients with AITL, and AMC >0.8 × 10(9)/L can routinely be used to identify high-risk patients with unfavorable survival.