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The highly heterogeneous characteristics of Wharton's jelly mesenchymal stem cells (WJ-MSCs) may be responsible for the poor clinical outcomes and poor reproducibility of treatments based on WJ-MSCs. Exploration of WJ-MSC heterogeneity with multimodal single-cell technologies will aid in establishing accurate MSC subtyping and developing screening protocols for dominant functional subpopulations. Here, the characteristics of WJ-MSCs are systematically analyzed by single cell and spatial transcriptome sequencing. Single-cell transcriptomics analysis identifies four WJ-MSC subpopulations, namely proliferative_MSCs, niche-supporting_MSCs, metabolism-related_MSCs and biofunctional-type_MSCs. Furthermore, the transcriptome, cellular heterogeneity, and cell-state trajectories of these subpopulations are characterized. Intriguingly, the biofunctional-type MSCs (marked by S100A9, CD29, and CD142) selected in this study exhibit promising wound repair properties in vitro and in vivo. Finally, by integrating omics data, it has been found that the S100A9

BACKGROUND: Lung cancer is the leading cause of cancer mortality in China. The clinicopathologic features and genetic profile of Chinese lung cancer patients need to be investigated. This study evaluated the gene mutation profile, analyzed the frequency of concurrent genes in epidermal growth factor receptor (EGFR)-mutant non-small cell lung cancer (NSCLC) patients, and determined its prognostic value. METHODS: We collected the clinical data from 151 initially diagnosed patients NSCLC. Tumor samples underwent targeted next-generation sequencing (NGS). Progression-free survival (PFS) and overall survival (OS) were analyzed using the Kaplan-Meier method. RESULTS: Among the 151 participants, the mutational profile revealed alterations in 29 genes, where TP53 (37.09%) and EGFR (30.46%) exhibited the highest mutation rates. Mutations in the EGFR gene were most prevalent (40%) in adenocarcinoma (LUAD) and were only present in 8.8% of participants with squamous cell carcinoma (LUSC). The most frequently mutated genes in LUAD patients were TP53 (47%), followed by KRAS (11.7%). In all 39 participants with EGFR mutations, TP53, KRAS, PIK3CA, APC, and FBXW7 were also mutated. Those with only EGFR mutation appeared to have a better prognosis; however, the difference was not statistically significant. Tumor mutational burden (TMB) was roughly significantly increased in patients who harbored EGFR and other mutant driver genes, compared with only EGFR mutant patients. The TMB value was significantly higher in those with P53 mutation than in P53 wild-type patients. CONCLUSIONS: We described the genetic profiles of NSCLC and compared the difference in genetic profiles between LUAD and LUSC. The concomitant genetic alterations might be a poor prognostic factor for patients with EGFR mutation, and TMB might be prognostically related to the co-mutations of EGFR and other genes.

Aim: The role of plasma heat shock protein 90 alpha (HSP90α) in colorectal cancer patients remains unclear. This study aimed to evaluate the relationship between HSP90α and the occurrence and development of colorectal cancer through diagnosis and prognosis value.Methods: 635 colorectal cancer patients and 295 healthy controls were recruited. The HSP90α was measured by using the ELISA kit in all objects and the immune cells and common biomarkers as CEA, AFP, CA125, CA153 and CA199 were measured in all colorectal cancer patients. The relationship between plasma HSP90α with clinical features, common tumor markers and immune cells were also conducted. The survival analysis endpoint was progression-free survival (PFS).Results: The levels of plasma HSP90α were significantly higher in colorectal cancer patients compared to healthy controls [51.4 (ng/ml) vs. 43.7 (ng/ml), p < 0.001]. In additional, the levels of plasma HSP90α were associated with gender and disease progress as stage, lymphatic and distant metastasis. Furthermore, plasma HSP90α was closed correlation with CEA, CA125, CA199 and percentage of B cells. However, the initial expression level of plasma HSP90α failed to show a prognostic value for progression-free survival in colorectal cancer.Conclusion: The plasma Hsp90α was remarkable higher in colorectal cancer and correlated with common tumor biomarkers and immune cells. Plasma Hsp90α levels were associated with disease progress but a poor diagnosis performance and also failed to show a prognostic value in colorectal cancer.

To investigate the distribution of class 1 integrons and their variable regional molecular characteristics, as well as the diversity of promoter and drug sensitivity of CR-Eco (carbapenem-resistant E. coli) strains.A total of 117 CR-Eco strains, collected between 2012.01 and 2019.12, underwent fully automated bacterial identification and sensitization using VITEK-2 Compact and supplemented by K-B assay. PCR was employed to screen for class 1 integrase genes and integron variable regions, while the promoter type and variable region gene cassette characteristics were determined by sequencing analysis.The positive rate of the class 1 integron of the CR-Eco strains was 83.70% (92/117) herein. Moreover, class 1 integrase-positive strains exhibited statistically significant resistance to aztreonam, ceftazidime, ciprofloxacin, ceftriaxone, gentamicin, meropenem, and trimethoprim-sulfamethoxazole compared to integron-negative strains (P 0.05). Variable regions were observed in 77 of the 92 class 1 integrase-positive strains. In addition, seven gene cassettes were detected, namely dfrA17-aadA5, aadA22, dfrA12-aadA2, dfrA12, dfrA17, dfrA27 and aadA. Finally, five types of class 1 integron variable region promoters were identified in those 77 strains, including PcW, PcH1, PcWThe primary integrator variable region gene cassettes of this class were dfrA and aadA. The integron-positive strains displayed simultaneous high resistance to multiple antimicrobial drugs. The integrator variable region promoters of the CR-Eco strains are primarily weak and can potentially form and spread drug resistance.

The main strategy of tissue repair and regeneration focuses on the application of mesenchymal stem cells and cell-based nanoparticles, but there are still multiple challenges that may have negative impacts on human safety and therapeutic efficacy. Cell-free nanotechnology can effectively overcome these obstacles and limitations. Mesenchymal stem cell (MSC)-derived natural small extracellular vesicles (sEVs) represent ideal nanotherapeutics due to their low immunogenicity and lack of tumorigenicity. Here, sEVs harvested from Wharton's jelly mesenchymal stem cells (WJMSCs) were identified. In vitro results showed that WJMSC-sEVs efficiently entered chondrocytes in the osteoarthritis (OA) model, further promoted chondrocyte migration and proliferation and modulated immune reactivity. In vivo, WJMSC-sEVs notably promoted chondrogenesis, which was consistent with the effect of WJMSCs. RNA sequencing results revealed that sEV-microRNA-regulated biocircuits can significantly contribute to the treatment of OA, such as by promoting the activation of the calcium signaling pathway, ECM-receptor interaction pathway and NOTCH signaling pathway. In particular, let-7e-5p, which is found in WJMSC-sEVs, was shown to be a potential core molecule for promoting cartilage regeneration by regulating the levels of STAT3 and IGF1R. Our findings suggest that WJMSC-sEV-induced chondrogenesis is a promising innovative and feasible cell-free nanotherapy for OA treatment.

Background Traditional therapies fail to cure most glioblastoma patients and the 5-year survival rate is less than 10%, highlighting need for new therapeutic approaches. The natural killer group 2 member D ligands (NKG2DLs) are highly expressed in glioblastomas and are considered promising targets for chimeric antigen receptor (CAR) T-cell therapy. The aim of this study was to investigate the effect of NKG2D-expressing CAR-T cells on glioblastomas and glioblastoma stem cells. Methods The expression of NKG2DLs was analyzed by flow cytometry and immunohistochemistry. NKG2D-BBz CAR, containing the extracellular domain of NKG2D, was constructed and delivered into T cells by lentiviral particles. In vitro cytotoxicity of the CAR-T cells was assessed by flow cytometry. Release of cytokine, perforin and granzyme B was quantified using enzyme-linked immunosorbent assay kits. The therapeutic efficacy of NKG2D-BBz CAR-T cells in vivo was evaluated using subcutaneous tumor models. The safety of the CAR was analyzed by investigating the effects on proliferation, apoptosis, and karyotype. Results Our data confirmed the high expression of NKG2DLs in human glioblastoma cells, cancer stem cells, and tumor samples. Further, the NKG2D-BBz CAR-T cells efficiently lysed glioblastoma cells and cancer stem cells in vitro and produced high levels of cytokines, perforin, and granzyme B. The CAR-T cells markedly eliminated xenograft tumors in vivo and did not exhibit significant treatment-related toxicity in the treated mice. The CAR expression also did not exert any obvious effects on cell proliferation, apoptosis, and genomic stability. Conclusion Our findings demonstrated that NKG2D CAR-T cells targeted glioblastoma cells and cancer stem cells in an NKG2D-dependent manner, supporting the use of CAR-T therapy in glioblastoma therapeutic strategies. Electronic supplementary material The online version of this article (10.1186/s40425-019-0642-9) contains supplementary material, which is available to authorized users.

In this paper, correspondence relations between the solutions of the static bending of functionally graded material (FGM) Reddy–Bickford beams (RBBs) and those of the corresponding homogenous Euler–Bernoulli beams are presented. The effective material properties of the FGM beams are assumed to vary continuously in the thickness direction. Governing equations for the titled problem are formulated via the principle of virtual displacements based on the third-order shear deformation beam theory, in which the higher-order shear force and bending moment are included. General solutions of the displacements and the stress resultants of the FGM RBBs are derived analytically in terms of the deflection of the reference homogenous Euler–Bernoulli beam with the same geometry, loadings and end conditions, which realize a classical and homogenized expression of the bending response of the shear deformable non-homogeneous FGM beams. Particular solutions for the FGM RBBs under specified end constraints and load conditions are given to validate the theory and methodology. The key merit of this work is to be capable of obtaining the high-accuracy solutions of thick FGM beams in terms of the classical beam theory solutions without dealing with the solution of the complicated coupling differential equations with boundary conditions of the problem.

Background Next generation sequencing revealed frequent mutations of NRAS, KRAS or BRAF in up to 50% of newly diagnosed multiple myeloma (MM) patients. Methods Specific K- or N-RAS knockdowns led to strong decrease in MM cell viability if harbored the respective oncogenic isoform, but minimal effects were observed if only the wild-type isoform was present, presenting RAS signaling as the key driving oncogenes in the mutation bearing patients. Unfortunately, no inhibitor for RAS mutation is available for clinical use in MM. Therefore, key component in the RAS/MAPK pathway may represent an alternative therapeutic target for MM. Germinal center kinase (GCK) is an upstream activator in the MAPK pathway. Results Our data showed that GCK (MAP4K2) knockdown in MM cells induced MM cell growth inhibition, associated with the downregulation of critical transcriptional factors including IKZF1/3, BCL-6, and c-MYC proteins. Moreover, GCK silencing only led to significantly decreased GCK mRNA, however, did not affect IKZF1 expressions at mRNA level. We next tested the effects of GCK inhibitor TL4-12 on MM. Given the fact that RASMut MM cells have higher GCK expression level and are more sensitive to GCK knockdown, we compared mutated RAS MM cells with wild-type RAS MM cells. TL4-12 significantly inhibited the growth of mutated RAS MM cells, with IC50 5-10 fold lower compared to wild-type RAS MM cell lines. IKZF1/3 are the key targets of the immunomodulatory drugs (IMiDs), which are the backbone of MM therapy. IMiDs bind to cereblon (CRBN) and induce IKZF1/3 protein degradation, which subsequently lead to MM cell growth inhibition. Our data showed that GCK knockdown also downregulates IKZF1 protein level, indicating that IKZF1 is under the regulation of GCK. Concomitant treatment with proteasome inhibitor PS-341 blocked TL4-12 induced IKZF1 downregulation, suggesting that inhibition of GCK induces IKZF1 protein degradation. However, since IMiDs-resistant RPMI-8226 and JJN3 MM cells are sensitive to GCK inhibition, we hypothesized that the IKZF1 degradation mechanism induced by GCK inhibition is different from IMiDs and independent of CRBN. To confirm this hypothesis, we silenced CRBN in N-Rasmut H929 MM cells and examined the response to IMiDs and GCK inhibitor. CRBN silencing in H929 cells resulted in lenalidomide resistance. In contrast, CRBN silencing failed to rescue H929 MM from TL4-12 induced proliferation inhibition and IKZF1 downregulation, confirming that GCK regulated IKZF1 and cell growth is independent of CRBN. Conclusions Taken together, our data demonstrated that GCK inhibition induces cell growth inhibition and triggers apoptosis especially in RASmut MM cells. Importantly, GCK inhibition downregulates IKZF1 via a CRBN-independent mechanism. Our findings thus provide a rationale for the clinical evaluation of targeting GCK in RASmut MM patients and further mechanistic insight into the role of GCK in MM tumorigenesis as well as drug resistance.

Subarachnoid hemorrhage (SAH) is a cerebrovascular disease associated with high morbidity and mortality. CXCR4 provides neuroprotective effects, which can alleviate brain injury and inflammation induced by stroke. Previous studies have suggested that CXCR4 reduces the pyroptosis of LPS-stimulated BV2 cells. The purpose of this study was to evaluate the antipyroptosis effects and mechanisms of CXCR4 after SAH. SAH animal model was induced via endovascular perforation. A total of 136 male Sprague-Dawley rats were used. Recombinant human cysteine-X-cysteine chemokine ligand 12 (rh-CXCL-12) was administered intranasally at 1 h after SAH induction. To investigate the underlying mechanism, the inhibitor of CXCR4, AMD3100, was administered intraperitoneally at 1 h before SAH. The neurobehavior tests were assessed, followed by performing Western blot and immunofluorescence staining. The Western blot results suggested that the expressions of endogenous CXCL-12, CXCR4, and NLRP1 were increased and peaked at 24 h following SAH. Immunofluorescence staining showed that CXCR4 was expressed on neurons, microglia, and astrocytes. Rh-CXCL-12 treatment improved the neurological deficits and reduced the number of FJC-positive cells, IL-18-positive neurons, and cleaved caspase-1(CC-1)-positive neurons after SAH. Meanwhile, rh-CXCL-12 treatment increased the levels of CXCL-12 and CXCR4, and reduced the levels of NLRP1, IL-18, IL-1β, and CC-1. Moreover, the administration of AMD3100 abolished antipyroptosis effects of CXCL-12 and its regulation of CXCR4 post-SAH. The CXCR4/NLRP1 signaling pathway may be involved in CXCL-12-mediated neuronal pyroptosis after SAH. Early administration of CXCL-12 may be a preventive and therapeutic strategy against brain injury after SAH.

Background Idiopathic pulmonary fibrosis (IPF) is a progressive disease resulting in respiratory failure with no efficient treatment options. We investigated the protective effect of RS4651 on pulmonary fibrosis in mice and the mechanism. Methods Intratracheal injection of bleomycin (BLM) was used to induce pulmonary fibrosis in mice. RS4561 was administered intraperitoneally at different doses. Histopathological changes were observed. The level of alpha-smooth muscle actin (α-SMA) were also tested. In vitro, the proliferation and migratory effects of RS4651 treatment on MRC-5 cells pre-treated with transforming growth factor (TGF-β1) were examined. RNA-sequencing was used to detect differentially expressed target genes. Then, the expression of α-SMA, pSMAD2 and SMAD7 were analysed during RS4651 treatment of MRC-5 cells with or without silencing by SMAD7 siRNA. Results Histopathological staining results showed decreased collagen deposition in RS4651 administered mice. Additionally, a lower level of α-SMA was also observed compared to the BLM group. The results of in vitro studies confirmed that RS4651 can inhibit the proliferation and migration, as well as α-SMA and pSMAD2 expression in MRC-5 cells treated with TGF-β1. RNA-sequencing data identified the target gene SMAD7. We found that RS4651 could upregulate SMAD7 expression and inhibit the proliferation and migration of MRC-5 cells via SMAD7, and RS4651 inhibition of α-SMA and pSMAD2 expression was blocked in SMAD7-siRNA MRC-5 cells. In vivo studies further confirmed that RS4651 could upregulate SMAD7 expression in BLM-induced lung fibrosis in mice. Conclusions Our data suggest that RS4651 alleviates BLM-induced pulmonary fibrosis in mice by inhibiting the TGF-β1/SMAD signalling pathway.

This is the first report of ceftazidime–avibactam resistance caused by the blaKPC-33 mutation through the D179Y variant during the treatment of blaKPC-2-positive Klebsiella pneumoniae-related infections in China. The blaKPC-33-containing K. pneumoniae was susceptible to meropenem–vaborbactam, cefepime–zidebactam, tigecycline, and polymyxin B. The blaKPC-33 gene was located on a 77 551-bp transformable plasmid harboring qnrS1 and blaLAP-2. Detecting blaKPC-33-positive K. pneumoniae clinical strains is important for infection control.

BackgroundSubarachnoid hemorrhage (SAH) is a cerebrovascular disease associated with high morbidity and mortality. CXCR4 provides a neuroprotective effect, which can alleviate brain injury and inflammation induced by stroke. The purpose of this study was to evaluate the anti-inflammatory effects and mechanisms of CXCR4 after SAH. Methods: SAH was induced via endovascular perforation. 185 male Sprague-Dawley rats were used. Recombinant human cysteine-X-cysteine chemokine ligand 12 (rh-CXCL-12) was administered intranasally at 1 h after SAH induction. To investigate the underlying mechanism, the inhibitors of CXCR4 and P13K, AMD3100 and LY294002, respectively, were administered intraperitoneally at 1 h before SAH. The short- and long-term neurobehavior were assessed, followed by performing western blot and immunofluorescence staining. ResultsWestern blotting suggested that the expressions of endogenous CXCL-12 and CXCR4 were increased, and peaked at 24 h following SAH. Immunofluorescence staining showed that CXCR4 was expressed on microglia. Rh-CXCL-12 treatment reduced the number of M1 macrophages and improved the short- and long-term neurological deficits after SAH. Meanwhile, rh-CXCL-12 treatment increased the levels of CXCL-12, CXCR4, PI3K, and p-Akt, and reduced the levels of IL-1β, IL-6, and TNF-α. Moreover, the administration of AMD3100 and LY294002 abolished the post-SAH neurobehavioral and neuroinflammatory improvements of CXCL-12 and its regulation of PI3K and p-Akt protein levels.Conclusions The CXCR4/PI3K/Akt signaling pathway may be involved in CXCL-12-mediated reduction of post-SAH neuroinflammation. Early administration of CXCL-12 may be a preventive and therapeutic strategy against delayed brain injury after SAH.

Background Mesenchymal stem cell-based acellular therapies have been widely exploited in managing hypertrophic scars. However, low maintenance dose and transitory therapeutic effects during topical medication remain a thorny issue. Herein, this study aimed to optimize the curative effect of adipose-derived stem cell conditioned medium (ADSC-CM) in the prevention of hypertrophic scarring. Methods In the present study, ADSC-CM was concentrated via the freeze-drying procedure. The efficacy of different dose groups (CM, CM5, CM10) was conducted on the proliferation, apoptosis, and α-smooth muscle actin (α-SMA) expression of human keloid fibroblasts (HKFs) in vitro. Incorporation of adipose-derived stem cell concentrated conditioned medium (ADSCC-CM) into polysaccharide hydrogel was investigated in rabbit ear, in vivo. Haematoxylin-eosin (H&E) and Masson’s trichrome staining were performed for the evaluation of scar hyperplasia. Results We noted that ADSCC-CM could downregulate the α-SMA expression of HKFs in a dose-dependent manner. In the rabbit ear model, the scar hyperplasia in the medium-dose group (CM5) and high-dose group (CM10) was inhibited with reduced scar elevation index (SEI) under 4 months of observation. It is noteworthy that the union of CM5 and polysaccharide hydrogel (CM5+H) yielded the best preventive effect on scar hyperplasia. Briefly, melanin, height, vascularity, and pliability in the CM5+H group were better than those of the control group. Collagen was evenly distributed, and skin appendages could be regenerated. Conclusions Altogether, ADSCC-CM can downregulate the expression of α-SMA due to its anti-fibrosis effect and promote the rearrangement of collagen fibres, which is integral to scar precaution. The in situ cross bonding of ADSCC-CM and polysaccharide hydrogel could remarkably enhance the therapeutic outcomes in inhibiting scar proliferation. Hence, the alliance of ADSCC-CM and hydrogel may become a potential alternative in hypertrophic scar prophylaxis.

Oxidative stress (OS) and neuronal apoptosis are major pathological processes after hypoxic-ischemic encephalopathy (HIE). Colony stimulating factor 1 (CSF1), binding to CSF1 receptor (CSF1R), has been shown to reduce neuronal loss after hypoxic-ischemia- (HI-) induced brain injury. In the present study, we hypothesized that CSF1 could alleviate OS-induced neuronal degeneration and apoptosis through the CSF1R/PLCG2/PKA/UCP2 signaling pathway in a rat model of HI. A total of 127 ten-day old Sprague Dawley rat pups were used. HI was induced by right common carotid artery ligation with subsequent exposure to hypoxia for 2.5 h. Exogenous recombinant human CSF1 (rh-CSF1) was administered intranasally at 1 h and 24 h after HI. The CSF1R inhibitor, BLZ945, or phospholipase C-gamma 2 (PLCG2) inhibitor, U73122, was injected intraperitoneally at 1 h before HI induction. Brain infarct volume measurement, cliff avoidance test, righting reflex test, double immunofluorescence staining, western blot assessment, 8-OHdG and MitoSOX staining, Fluoro-Jade C staining, and TUNEL staining were used. Our results indicated that the expressions of endogenous CSF1, CSF1R, p-CSF1R, p-PLCG2, p-PKA, and uncoupling protein2 (UCP2) were increased after HI. CSF1 and CSF1R were expressed in neurons and astrocytes. Rh-CSF1 treatment significantly attenuated neurological deficits, infarct volume, OS, neuronal apoptosis, and degeneration at 48 h after HI. Moreover, activation of CSF1R by rh-CSF1 significantly increased the brain tissue expressions of p-PLCG2, p-PKA, UCP2, and Bcl2/Bax ratio, but reduced the expression of cleaved caspase-3. The neuroprotective effects of rh-CSF1 were abolished by BLZ945 or U73122. These results suggested that rh-CSF1 treatment attenuated OS-induced neuronal degeneration and apoptosis after HI, at least in part, through the CSF1R/PLCG2/PKA/UCP2 signaling pathway. Rh-CSF1 may serve as therapeutic strategy against brain damage in patients with HIE.

Background Hypoxic-ischemic encephalopathy (HIE) is a life-threatening cerebrovascular disease. Neuroinflammation plays an important role in the pathogenesis of HIE, in which microglia are key cellular mediators in the regulation of neuroinflammatory processes. Colony-stimulating factor 1 (CSF1), a specific endogenous ligand of CSF1 receptor (CSF1R), is crucial in microglial growth, differentiation, and proliferation. Recent studies showed that the activation of CSF1R with CSF1 exerted anti-inflammatory effects in a variety of nervous system diseases. This study aimed to investigate the anti-inflammatory effects of recombinant human CSF1 (rh-CSF1) and the underlying mechanisms in a rat model of HIE. Methods A total of 202 10-day old Sprague Dawley rat pups were used. HI was induced by the right common carotid artery ligation with subsequent exposure of 2.5-h hypoxia. At 1 h and 24 h after HI induction, exogenous rh-CSF1 was administered intranasally. To explore the underlying mechanism, CSF1R inhibitor, BLZ945, and phospholipase C-gamma 2 (PLCG2) inhibitor, U73122, were injected intraperitoneally at 1 h before HI induction, respectively. Brain infarct area, brain water content, neurobehavioral tests, western blot, and immunofluorescence staining were performed. Results The expressions of endogenous CSF1, CSF1R, PLCG2, protein kinase C epsilon type (PKCε), and cAMP response element-binding protein (CREB) were gradually increased after HIE. Rh-CSF1 significantly improved the neurological deficits at 48 h and 4 weeks after HI, which was accompanied by a reduction in the brain infarct area, brain edema, brain atrophy, and neuroinflammation. Moreover, activation of CSF1R by rh-CSF1 significantly increased the expressions of p-PLCG2, p-PKCε, and p-CREB, but inhibited the activation of neutrophil infiltration, and downregulated the expressions of IL-1β and TNF-α. Inhibition of CSF1R and PLCG2 abolished these neuroprotective effects of rh-CSF1 after HI. Conclusions Our findings demonstrated that the activation of CSF1R by rh-CSF1 attenuated neuroinflammation and improved neurological deficits after HI. The anti-inflammatory effects of rh-CSF1 partially acted through activating the CSF1R/PLCG2/PKCε/CREB signaling pathway after HI. These results suggest that rh-CSF1 may serve as a potential therapeutic approach to ameliorate injury in HIE patients.

Objectives To assess the efficacy of corticosteroid treatment of patients with coronavirus disease 2019 (COVID-19). Design, setting Observational study in the two COVID-19-designated hospitals in Wuhu, Anhui province, China, 24 January - 24 February 2020. Participants Thirty-one patients infected with the severe acute respiratory coronavirus 2 (SARS-CoV-2) treated at the two designated hospitals. Main outcome measures Virus clearance time, length of hospital stay, and duration of symptoms, by treatment type (including or not including corticosteroid therapy). Results Eleven of 31 patients with COVID-19 received corticosteroid treatment. Cox proportional hazards regression analysis indicated no association between corticosteroid treatment and virus clearance time (hazard ratio [HR], 1.26; 95% CI, 0.58-2.74), hospital length of stay (HR, 0.77; 95% CI, 0.33-1.78), or duration of symptoms (HR, 0.86; 95% CI, 0.40-1.83). Univariate analysis indicated that virus clearance was slower in two patients with chronic hepatitis B infections (mean difference, 10.6 days; 95% CI, 6.2-15.1 days). Conclusions Corticosteroids are widely used when treating patients with COVID-19, but we found no association between therapy and outcomes in patients without acute respiratory distress syndrome. An existing HBV infection may delay SARS-CoV-2 clearance, and this association should be further investigated.

OBJECTIVE. This study aims to assess correlations of the time from symptom onset to diagnosis and treatment with the time to disease resolution and CT scores as based on findings from sequential chest CT examinations. MATERIALS AND METHODS. Thirty patients with coronavirus disease (COVID-19) confirmed by reverse transcription-polymerase chain reaction analysis underwent chest CT examinations. Five patients who did not have positive CT findings or who had not yet fulfilled criteria for discharge from the hospital were excluded. CT scores were determined according to CT findings and lung involvement. The time from symptom onset to diagnosis and treatment was recorded for each patient, and on the basis of this information, patients with COVID-19 were divided into group 1 (patients for whom this interval was ≤ 3 days) and group 2 (those for whom this interval was > 3 days). The CT scores for each group were fitted using a Lorentzian line-shape curve to show the variation tendency during treatment. The differences in age, sex, and last CT scores determined before discharge between the two groups were analyzed, and correlations of the time from symptom onset to diagnosis and treatment with the time to disease resolution as well as with the highest CT score also underwent statistical analysis. RESULTS. A total of 25 subjects were enrolled in the study. The fitted tendency curves for group 1 and group 2 were significantly different, with peak points showing that the estimated highest CT score was 10 and 16 for each group, respectively, and the time to disease resolution was 6 and 13 days, respectively. The Mann-Whitney test showed that the last CT scores were lower for group 1 than for group 2 (p = 0.025), although the chi-square test found no difference in age and sex between the groups. The time from symptom onset to diagnosis and treatment had a positive correlation with the time to disease resolution (r = 0.93; p = 0.000) as well as with the highest CT score (r = 0.83; p = 0.006). CONCLUSION. Timely diagnosis and treatment are key to providing a better prognosis for patients with COVID-19.

Objective. The focus of the present study is to evaluate the effects of Angiotensin II (Ang II) on neuronal apoptosis after HIE and the potential underlying mechanisms. Methods. Primary neonatal rat cortical neurons were used to study the oxygen-glucose deprivation (OGD) cell model. The expressions of Ang II, AT1R, GSK-3β, p-GSK-3β, mTOR, p-mTOR, Bax, Bcl-2, and cleaved caspase-3 were detected via western blot. IF and flow cytometry were used to evaluate neuronal apoptosis. Hypoxic-ischemic encephalopathy (HIE) was established to evaluate the therapeutic effects of Ang II in vivo. Cerebral infarction areas were detected by 2,3,5-Triphenyltetrazolium chloride staining. The righting and geotaxis reflexes were also recorded. In addition, Fluoro-Jade C staining and TUNEL staining were performed to evaluate neuronal degeneration and apoptosis. Results. Ang II significantly increased the rate of neuronal apoptosis, upregulated the expression of cleaved caspase-3, and downregulated Bcl-2/Bax ratio after OGD insult. For vivo assay, the expressions of endogenous Ang II and AT1R gradually increased and peaked at 24 h after HIE. Ang II increased NeuN-positive AT1R cell expression. In addition, Ang II increased the area of cerebral infarction, promoted neuronal degeneration and apoptosis, aggravated neurological deficits on righting and geotaxis reflexes, and was accompanied by increased expressions of phosphorylated GSK-3β and mTOR. The application of valsartan (Ang II inhibitor) or SB216763 (GSK-3β inhibitor) reversed these phenomena triggered by Ang II following HIE. Conclusion. Ang II increased neuronal apoptosis through the AT1R/GSK-3β/mTOR signaling pathway after experimental HIE both in vitro and in vivo, and Ang II may serve as a novel therapeutic target to ameliorate brain injury after HIE.

The effects of XCHD on the proliferation of C6 cells and on factors associated with the microRNA-34a (miR-34a)/p53/caspase-3 signaling pathway in vitro were investigated. Methods. XCHD was purchased too much to complete the study. CCK-8 assay was used to measure the XCHD concentration, and qPCR was used to quantify miR-34a expression at the mRNA level. Apoptosis was assessed using TUNEL. Western blots were used to determine the p53, caspase-3, caspase-8, and Bcl-2 expression levels. Results. The optimal XCHD concentration and time effect for C6 cells were observed after 36 h of exposure to a concentration of 100 µg/ml XCHD. miR-34a expression increased 8 and 12 h after the addition of XCHD. The presence of XCHD decreased Bcl-2 expression but increased p53, cleaved caspase-3, Bax, and caspase-8 expression. When p53 was inhibited, miR-34a expression was unaffected by the addition of XCHD, Bcl-2 expression was low, and cleaved caspase-3, Bax, and caspase-8 expression increased. The inhibition of p53 promoted C6 cell growth when compared with C6 cells exposed to XCHD and with no inhibition of p53. Conclusions. XCHD inhibits C6 cell growth which was influenced by the p53/caspase pathway.

The immunomodulatory drugs, lenalidomide and pomalidomide yield high response rates in multiple myeloma patients, but are associated with a high rate of thrombocytopenia and increased risk of secondary hematologic malignancies. Here, we demonstrate that the immunomodulatory drugs induce self-renewal of hematopoietic progenitors and upregulate megakaryocytic colonies by inhibiting apoptosis and increasing proliferation of early megakaryocytic progenitors via down-regulation of IKZF1. In this process, the immunomodulatory drugs degrade IKZF1 and subsequently down-regulate its binding partner, GATA1. This results in the decrease of GATA1 targets such as ZFPM1 and NFE2, leading to expansion of megakaryocytic progenitors with concomitant inhibition of maturation of megakaryocytes. The down-regulation of GATA1 further decreases CCND1 and increases CDKN2A expression. Overexpression of GATA1 abrogated the effects of the immunomodulatory drugs and restored maturation of megakaryocytic progenitors. Our data not only provide the mechanism for the immunomodulatory drugs induced thrombocytopenia but also help to explain the higher risk of secondary malignancies and long-term cytopenia induced by enhanced cell cycling and subsequent exhaustion of the stem cell pool.

Full-thickness skin damage is a server issue and sometimes even dangerous to life. Many researches have been done toward full-thickness wound dressing. In this study, we demonstrated a facile and one-step procedure of SIS bilayer wound dressing. The top layer could protect the wound from bacterial infection and provide a moist environment suitable for wound healing, while the cryogel layer could promote cell proliferation. The SIS bilayer wound dressing has sufficient mechanical properties to protect wound from second damage and can maintain a moist environment for cell proliferation and migration at wound site. Bacterial permeation testing demonstrated that the bilayer scaffold had high efficiency in blocking bacteria at the wound site. In vivo tests and qRT-PCR results revealed that the bilayer group possessed a higher tendency toward keratinocyte proliferation and migration. The SIS bilayer has a high potential to use as full-thickness wound dressing.

Introduction Multiple myeloma (MM)-induced bone disease remains one of its most devastating complications, caused by increased bone resorption by overactivated osteoclasts coupled with impaired bone formation. MM cells produce osteoclast-activating factors that induce osteoclast activation and extensive bone resorption. Our previous work demonstrated that matrix metalloproteinase 13 (MMP-13) is a critical osteoclastogenic factor that is highly secreted by MM cells (Fu J etc. JCI. 2016). We also identified that the checkpoint inhibitor, programmed death-1 homolog (PD-1H/VISTA), serves as the MMP-13 receptor in osteoclasts and mediates MMP-13-dependent osteoclastogenic function which is largely blocked in Pd-1h -/-osteoclasts (Fu J etc. ASH 2019, 2020). While the inhibitory role of PD-1H/VISTA in T-cells has recently been described (ElTanbouly MA etc. Science 2020), its cellular binding proteins remain unclear, and its role in osteoclast activation andMM bone disease have not been addressed. Methods and Results To identify its interacting proteins, PD-1H-His 6 recombinant protein was expressed in mouse bone marrow mononuclear cells, the associated proteins pulled down by Ni-NTA agarose beads from cell lysates and identified by mass spectrometry. Functional annotation charting of the 75 proteins enriched in PD-1H pull-down samples (with signal ratio of PD-1H-His 6 pull-down vs control >2) indicated that almost 30% of the interacting targets were either cytoskeletal or cytoskeleton-associated proteins. Given that the F-actin cytoskeleton undergoes dynamic reorganization during osteoclast differentiation and plays critical roles in bone resorption, we further addressed the role of PD-1H in F-actin cytoskeleton regulation. Initially, osteoclasts form F-actin-rich adhesive structures, termed podosomes. At later stages, podosomes collectively rearrange into clusters and rings and finally into sealing belts to mediate osteoclast spreading, migration and bone resorption (Teitelbaum SL. Ann N Y Acad Sci. 2011). By confocal immunofluorescence microscopy, we found that PD-1H co-localized with F-actin podosome clusters, rings and sealing belts during osteoclast differentiation (Figure 1A). The functional role of PD-1H in F-actin cytoskeleton reorganization was addressed using Pd-1h -/- osteoclast wherein Pd-1h knockout lead to the disruption of podosome clusters at early stages relative to WT controls, while at later stages, Pd-1h -/-osteoclasts exhibited significantly fewer F-actin rings and belts (Figure 1B). Further, binding of MMP-13 to PD-1H increased the number of osteoclasts forming F-actin rings and belts, as well as the size of F-actin belts, which was blocked in Pd-1h -/- osteoclasts. To determine the role of PD-1H in the development of myeloma-induced lytic bone lesions, 5TGM1 myeloma cells were bilaterally intratibially injected into Pd-1h wtRag2 -/- or Pd-1h -/-Rag2 -/- mice (n=10) to induce lytic bone lesions. Three weeks following intratibial 5TGM1 injection, tibiae were harvested for micro-computed tomography. Subsequent quantitative histomorphology analyses of the trabecular and cortical bones confirmed that the knockout of Pd-1h reduced MM-induced bone destruction with significantly less decrease in trabecular bone volume (Tb. [BV/TV]), trabecular bone number (Tb. N.), trabecular bone thickness (Tb. Th.), as well as less increase in trabecular bone spacing (Tb. Sp.) and bone specific surface (Tb. [BS/BV]) compared to Pd-1h wtRag2 -/- mice. Similar effects were observed in cortical bone with less decrease in cortical bone thickness (CT. Th.), cortical bone area fraction (CT. [BA/TA]), and cortical tissue mineral density (CT. TMD) in 5TGM1 bearing Pd-1h -/-Rag2 -/- mice vs Pd-1h wtRag2 -/- mice (Table 1). Conclusions Taken together, our study, for the first time, reveals the novel role of checkpoint inhibitor, PD-1H/VISTA, in osteoclasts and myeloma bone disease. PD-1H associates with cytoskeleton proteins and regulates the F-actin cytoskeleton reorganization which is critical for osteoclast bone resorption activity. Further, PD-1H mediates MMP-13-induced osteoclast fusion, F-actin belts formation, and osteoclast activation. Pd-1h -/-in recipient mice significantly impairs MM-induced bone loss, demonstrating that PD-1H/VISTA plays a critical role in MM bone disease. Figure 1 Figure 1. Disclosures No relevant conflicts of interest to declare.

Recently, mitogen-activated protein kinase kinase kinase kinase 2 (MAP4K2) has emerged as an important key regulator of the stress-activated MAPK core signaling pathways. MAP4K2, also called Germinal Center Kinase (GCK), is predominantly and highly expressed in the germinal center of B cells. Recently we have shown that MAP4K2 knockdown in K- or N-RAS mutated MM cells induces MM cell growth inhibition, associated with the downregulation of critical transcriptional factors including IKZF1/3, BCL-6, and c-MYC proteins (Li et al. Blood 2021). Importantly, MAP4K2 silencing induces IKZF1 protein degradation without affecting IKZF1 mRNA level. Furthermore, IMiDs-resistant K-RAS Mut MM cells are sensitive to MAP4K2 inhibition induced IKZF1 degradation and cell growth suppression, suggesting that MAP4K2 inhibition overcomes IMiDs resistances in MM. To further validate MAP4K2 inhibition as a novel strategy to overcome IMiDs-resistance, we generated lenalidomide-resistant human myeloma cell lines. In this model, MM1S-LEN RES cells showed significantly decreased expression of CRBN protein compared to the parent cells. Accordingly, upon lenalidomide treatment, CRBN-mediated down-regulation of IKZF1, c-MYC, IKZF3 and IRF4 were abrogated in the MM1S-LEN RES cells. As expected, MM1S-LEN RES cells were resistant to lenalidomide induced growth inhibition in cell proliferation assay. In contrast, MAP4K2 inhibition using TL4-12 potently induced IKZF1, c-MYC, and IRF4 downregulation as well as cell proliferation inhibition, demonstrating that MAP4K2 regulates IKZF1 and cell growth independently of CRBN. These results indicate that MAP4K2 is a novel therapeutic target to overcome IMiDs-resistance MM. Iberdomide (CC-220) is an orally available IMiD® compound under development for the treatment of relapsed/refractory multiple myeloma. Previous biochemical and structural studies demonstrated that Iberdomide binds to cereblon with a higher affinity than lenalidomide or pomalidomide. Here, we evaluated the combination effects of Iberdomide with MAP4K2 silencing in MM. Tet-on sh-MAP4K2 lentivirus were introduced into RAS Mut MM cells to establish the inducible MAP4K2 knockdown cells upon doxycycline treatment. To address the combination effects, Tet-on sh-MAP4K2 RAS Mut MM cells were treated with doxycycline and different dosages of iberdomide. We found that MAP4K2 silencing strongly increased iberdomide-induced apoptosis (iberdomide alone vs. with MAP4K2 KD: 32% vs 92). Similar, in western blot assays, MAP4K2 silencing combined with Iberdomide significantly enhanced downregulation of IKZF1, c-MYC, and IRF4 compared to the iberdomide treatment alone. These data suggest that combination of iberdomide and MAP4K2 inhibition have synergetic anti-MM effects. Taken together, our findings demonstrate that MAP4K2 is a novel therapeutic target to bypass IMiDs resistance in RAS mutated MM. Combination of MAP4K2 inhibition with Iberdomide results in synergetic anti-cancer effects in MM, therefore could be a potent novel therapeutic regimen for patients with relapsed/refractory multiple myeloma. Disclosures Marcireau: Sanofi: Current Employment.

Background Multiple myeloma (MM) bone disease remains one of the most devastating complications of this incurable cancer, causing bone fractures, pain, mobility issues, and neurological deficits. MM cells secret pro-osteoclastogenic factors which lead to osteoclast (OCL) activation. Our previous work showed that matrix metalloproteinase 13 (MMP-13) is a critical osteoclastogenic factor highly secreted by MM cells and induces OCL fusion and bone resorption independently of its proteolytic activity (JCI 2016). We recently reported that MMP-13 binds to checkpoint inhibitor programmed death-1 homolog (PD-1H/VISTA), a surface receptor that is expressed on OCLs and mediates MMP-13 induced OCL fusion and bone resorption activity (ASH 2019). Bone resorption activity is significantly impaired in Pd-1h-/- OCLs in vitro. However, the function of PD-1H in MM bone disease has not been defined. Methods The role of PD-1H in MM bone disease was investigated using the intratibial 5TGM1 Rag2-/- MM bone disease mice model. Pd-1h-/-Rag2-/- mice were generated by crossbreeding Pd-1h-/- with Rag2-/- mice. Firefly luciferase-expressing 5TGM1 cells were intratibially injected into age and sex-paired Rag2-/- or Pd-1h-/-Rag2-/- mice (N=10). 3 weeks later, tibiae were harvested for quantitative micro-CT followed by histological analysis. Results Morphological analyses of trabecular and cortical bones confirmed that Pd-1h-/- recipient mice exhibited significantly less 5TGM1-induced bone loss (P Conclusions Taken together, our study, for the first time, reveals that checkpoint inhibitor PD-1H/VISTA is the critical receptor for MMP-13 in osteoclasts, thereby mediating MMP-13-induced osteoclast fusion, activation, and bone resorption. MM-induced trabecular bone loss was significantly lower in Pd-1h-/- mice, demonstrating that PD-1H/VISTA plays a critical role in MMP-13-induced MM bone disease. Given the checkpoint role of PD-1H/VISTA in cancer immunosuppression, we further posit that targeting the interaction of MMP-13 and PD-1H may represent a novel therapeutic strategy to treat MM bone disease and modulate the MM immune environment.

Introduction Multiple myeloma (MM) bone disease remains one of the most devastating complications of this incurable cancer, causing bone fractures, pain, mobility issues and neurological deficits. MM cells produce osteoclast-activating factors that induce osteoclast activation, thereby leading to excessive bone resorption and lytic bone lesions1. Our previous work demonstrated that matrix metalloproteinase 13 (MMP-13) is a critical osteoclastogenic factor that is highly secreted by MM cells. MMP-13 induces osteoclast fusion and bone-resorption via a mechanism independent of its proteolytic activity2. We recently reported that MMP-13 binds to checkpoint inhibitor programmed death-1 homolog (PD-1H/VISTA), a surface receptor that is expressed in osteoclasts at high levels3. Binding of MMP-13 to PD-1H/VISTA induces osteoclast fusion and bone resorption activity whereas knockdown or knockout of PD-1H/VISTA largely block MMP-13 mediated effects on osteoclasts3. However, the function of PD-1H inMM bone disease in vitro or in vivo has not been previously defined. Methods and Results To confirm the role of PD-1H in MMP-13 induced bone disease in MM, we first conducted MM-osteoclast trans-well co-culture assay using murine MM cell line, 5TGM1 cells, and bone marrow mononuclear cells from Pd-1h-/- or wild type (WT) mice. 5TGM1 control cells or MMP-13 knockdown 5TGM1 cells were seeded in the upper wells of the transwell plates; while WT or Pd-1h-/- bone marrow mononuclear cells were seeded in the lower wells and cultured for osteoclast differentiation assessed by TRAP staining. Results show that 5TGM1 induced differentiation of WT osteoclasts with significantly increased osteoclast size and nuclei number/osteoclast. Consistent with our previous results2, MMP-13 knockdown blocked the 5TGM1 MM cells-induced activation of WT osteoclasts. In contrast, neither 5TGM1 MM cells nor MMP-13 knockdown cells had significant effects on Pd-1h-/- osteoclasts. Hence, knockout of Pd-1h abrogated MMP-13 mediated MM induction of osteoclasts, indicating that MMP-13/PD-1H interactions are critically involved in MM-induced osteoclast activation. The in vivo role of PD-1H in MM bone disease was investigated using the intratibial 5TGM1 Rag2-/- MM bone disease mice model2. For this purpose, Pd-1h-/-Rag2-/- mice were generated by crossbreeding C57BL/6 Pd-1h-/- with C57BL/6 Rag2-/- mice. 3x105 firefly luciferase expressing 5TGM1 cells (5TGM1-luc) were intratibially injected into age and sex-paired Rag2-/- or Pd-1h-/-Rag2-/- mice (N=5). Tumor progression was monitored by weekly bioluminescence imaging (BLI). 3 weeks after tumor inoculation, tibiae were harvested for quantitative micro-CT, followed by histological analysis. Histological staining showed that intratibial injection of 5TGM1-luc MM cells induced extensive lytic lesions and trabecular bone loss in Rag2-/- mice. In contrast, in Pd-1h-/-Rag2-/- mice,the bone structure was maintained with markedly less bone loss. Morphological analyses of trabecular bone across proximal tibiae further indicated that in Rag2-/- mice, 5TGM1 induced significant changes in bone microarchitecture, with decreased bone volume fraction (bone volume/tissue volume), connective density, trabecular bone numbers, and trabecular bone thickness, as well as increased trabecular bone spacing (Table 1). In contrast, in Pd-1h-/-Rag2-/- mice, 5TGM1 failed to induce significant loss of trabecular bone, confirming the critical role of PD-1H in MM induced bone disease in vivo. Conclusions Taken together, our study, for the first time, reveal that checkpoint inhibitor PD-1H/VISTA is the critical receptor for MMP-13 in osteoclasts, thereby mediating MMP-13-induced osteoclast fusion, activation and bone resorption. MM-induced trabecular bone loss was significantly lower in Pd-1h-/-mice, demonstrating that PD-1H/VISTA plays a critical role in MMP-13-induced MM bone disease. Given the checkpoint role of PD-1H/VISTA in cancer immunosuppression, we further posit that targeting the interaction of MMP-13 and PD-1H may represent a novel therapeutic strategy to treat MM bone disease and modulate the MM immune environment. References 1. Marino S, Petrusca DN, Roodman GD. Br J Pharmacol. 2019;10.1111/bph.14889. 2. Fu J, Li S, Feng R, et al. J Clin Invest. 2016;126(5):1759-1772. 3. Fu J, Li S, Yang C, et al. Blood. 2019; 134 (Supplement_1): 3072. Disclosures Lentzsch: Caelum Biosciences: Current equity holder in private company, Membership on an entity's Board of Directors or advisory committees; Janssen: Consultancy; Celularity: Consultancy, Other; Magenta: Current equity holder in private company; Karyopharm: Research Funding; Mesoblast: Divested equity in a private or publicly-traded company in the past 24 months.

8027 Background: Activating mutations of the RAS genes NRAS, KRAS, and HRAS ( RASmut) occur in up to 50% of MM and portend poor survival and high recurrence rates. MM cells with RASmut are susceptible to inhibition of germinal center kinase (GCK), resulting in IKAROS degradation independent of cereblon (CRBN). Selinexor is a selective inhibitor of nuclear export (SINE) compound that can induce IKAROS degradation through CRBN-independent pathways to overcome immunomodulatory drug resistance. We explored the benefit of selinexor treatment for pts with RASmut MM. Methods: In the randomized BOSTON study, pts with MM after 1-3 therapies received weekly XVd or twice weekly Vd. In the single-arm STORM study pts with penta-treated, triple class refractory MM were treated with twice weekly Xd. Both treatment regimens are now FDA approved. Mutations were assessed post-hoc by exome sequencing of 119 and 52 pts from BOSTON and STORM, respectively. Pts were considered RASmut if their MM had NRAS, KRAS or HRAS mutations in codons 12, 13 or 61. Results: There were 54 pts (45%) with RASmut in BOSTON (XVd = 26, Vd = 28), and 17 (33%) in STORM. In BOSTON, pts with RASmut MM treated with XVd had significantly longer progression-free survival (PFS) than those treated with Vd (median [med] = 12.9 vs 6.7 months [mo], hazard ratio [HR] = 0.48 [95% CI 0.24-0.97], p = 0.039). For pts treated with Vd, those with RASmut had significantly shorter overall survival (OS) compared to RASwild-type (WT) (med = 16.8 mo vs not reached [NR], HR = 2.87 [95% CI 1.03-7.99], p = 0.035). PFS was not significantly different (med = 6.74 vs 9.82 mo, HR = 1.64 [95% CI 0.88-3.07], p = 0.122). Amongst pts on XVd, there was no difference in survival between RASmut and RASWT pts (PFS: med = 12.8 vs 12.9 mo, HR = 1.08 [95% CI 0.52-2.26], p = 0.83; OS: med = NR vs NR, HR = 0.94 [95% CI 0.36-2.45], p = 0.91). In STORM, pts with RASmut had shorter OS compared to RASwt pts (med = 6.1 vs NR, HR = 2.05 [95% CI 1.22-5.19], p = 0.010). Preliminary studies to explore the mechanisms of action related to RASmut MM sensitivity to XVd demonstrated that selinexor treatment in vitro leads to downregulation of GCK. Conclusions: Despite typically having the worst outcomes, pts with RASmut MM had a similar benefit from XVd as RASWT MM, showing that the XVd combination can overcome RASmut. Mechanistically, selinexor induced down regulation of GCK and enhanced killing of RASmut MM cells. With a manageable safety profile, the XVd regimen could provide a viable treatment option to improve survival of pts with MM with RAS mutations. Clinical trial information: NCT03110562. [Table: see text]

An improved apparatus for measuring freezing points has been developed. Compared to the traditional Beckmann freezing point instrument, the improved one overcame prior difficulties with solidification of liquid and made the solid–liquid equilibrium reversible with heat compensation from a heating tube. The reliability and accuracy were carefully examined by determining the freezing point of the solvent and solution, as well as calculating the molar mass of the solute. It was found that the molar mass was in good agreement with the literature data. Thus, this improved device can effectively and successfully be used in undergraduate physical chemistry laboratory courses.

Suppose that G is a finite group and H is a subgroup of G. We say that H is a BNA-subgroup of G if either H x = H or x ∈ for all x ∈ G. The BNA-subgroups of G are between normal and abnormal subgroups of G. We obtain some new characterizations for finite groups based on the assumption that all cyclic subgroups are BNA-subgroups.

Glioblastoma multiform (GBM) is a highly aggressive brain tumor with poor life expectancy, and glioma stem cells (GSCs) are a small population of tumor cells existed in GBM, in which GSCs response to drive GBM recurrence, invasion and contribute to the anti-cancer resistance. GSCs have been identified and developed as a therapeutic target for GBM and can be used in drugs screening. Isocostunolide is a natural sesquiterpenoid and contained abundant resource in medicinal plants, but the anti-cancer efficacies of it against GSCs are still unexplored. In this investigation, the anti-tumor activity of isocostunolide against GSCs was investigated and the result demonstrated that it inhibited the growth of GSCs (GSC-3#, GSC-12#, GSC-18#) significantly with an IC50 value of 2.80μg/ml, 2.61μg/ml, 1.07μg/ml, respectively. In further mechanism study, isocostunolide inhibited GSCs cell proliferation, induced GSCs apoptosis significantly, as well as increased the proportion of the cleavage of caspase-3. The result suggested that isocostunolide induced GSCs apoptosis via the caspase dependent apoptotic pathway. Moreover, isocostunolide damaged GSCs colony formation capacity significantly and exhibited the anti-cancer efficacy against GSCs in vitro.

Background The population characteristics of Inflammatory Bowel Disease (IBD) in China are different from those of western countries, and this population lacks large sample clinical data. This study aimed to analyze the clinical characteristics of inpatients with severe complications of IBD in our center. Methods In this study, medical records of 510 hospitalized IBD patients (excluding pediatrics) were included, digestive tract perforation, massive digestive tract hemorrhage, thrombosis, toxic megacolon, digestive tract fistula, digestive tract stenosis, severe malnutrition, severe infection, and carcinogenesis were defined as severe complications. The clinical process and follow-up were retrospectively analyzed. Results The incidence of severe complications in patients with IBD was 39.02%, 71.31%, 23.81% and 50.68% for CD, UC and IBDU, respectively, and the incidence of severe complications in CD was significantly higher than that in UC (P10 years (χ2=4.054, P=0.044). The incidence of carcinogenesis in UC patients with the course of 0-10 years was 1.56%, which was significantly lower than 8.47% of UC patients with the course of >10 years (P for 10 years is an independent risk factor for carcinogenesis of UC patients.

Transcription factors (TFs) are convergence points of signaling cascades that coordinate cell differentiation, proliferation, survival, and migration; and are commonly deregulated in solid and hematologic malignancies, including multiple myeloma (MM). Several recent studies indicate that the inhibition of TFs may lead to selective tumor cell death with little or no consequences for normal cells due to redundancy in signaling pathways. Nuclear hormone receptor (NHR)- TFs belong to the most common therapies in oncology today. In contrast, non-NHR-TFs have been considered 'un- druggable' until most recently.This review article summarizes advances of our knowledge on the complex composition of non-NHR-TFs and their binding to cognate DNA sequences that are propelling the development of novel strategies in MM.Protein-protein and protein-DNA- binding inhibitors, proteolysis- targeting chimeric molecules, and chromatin remodeling/epigenetic reader inhibitors are among the most promising novel compounds with a potentially high therapeutic index; they are likely to once more advance MM treatment strategies and improve patient outcome in the near future.

Objective: To investigate the role of lncRNA Rpph1 on amyloid-β induced neuronal injury in SK-N-SH cells and underlying mechanism. Methods:In vitro Alzheimer’s disease (AD) model was established using the SK-N-SH cells treated with Aβ25-35 peptide. APPswe/PS1ΔE9 double transgenic mice were used as AD animal model. Rpph1 was over-expressed and miR-122 was inhibited or overexpressed in SK-N-SH cells via transfection with pcDNA3.1-oe Rpph1 vector, miR-122 inhibitor or miR-122 mimic, respectively. Cell viabilities and apoptosis were evaluated using MTT or flow cytometry assay, respectively. Quantitative real-time PCR (RT-qPCR) was used to determine expression of Rpph1 and miR-122. Western blotting was used to determine the expression of apoptosis related proteins as well as Wnt/β-catenin signaling related proteins. Dual luciferase reporter assay was conducted to confirm the binding of miR-122 with predictive binding site in 3’ UTR of Rpph1 and Wnt1. Results: Both lncRNA Rpph1 and miR-122 were up-regulated in AD mouse. Either over-expression of Rpph1 or inhibition of miR-122 restored the cell viability or decreased cell apoptosis rate in Aβ induced SK-N-SH cells. Overexpression of miR-122 inhibited the cell viability while did not influence the Aβ level in SK-N-SH cells. Furthermore, over-expression of Rpph1, as well as inhibition of miR-122, elevated Bcl-2, c-myc, Survivin and decreased Bax expression via activating Wnt/β-catenin signaling. Dual luciferase reporter assay showed that miR-122 could directly target to 3’UTR of Rpph1 and Wnt1. Conclusion: Both lncRNA Rpph1 and miR-122 were up-regulated in AD mouse and Rpph1 activated Wnt/β-catenin signaling to ameliorate amyloid-β induced neuronal apoptosis in SK-N-SH cells via direct targeting miR-122.

Figure S1. ULBP1 staining in a tissue microarray containing 60 glioblastoma tissues and 10 normal tissues, scale bar = 250 μm. Figure S2. ULBP3 staining in a tissue microarray containing 60 glioblastoma tissues and 10 normal tissues, scale bar = 250 μm. Figure S3. The cell-surface expression of CD3 in the indicated cells was analyzed by flow cytometry. The RAJI cell line was used as a negative control. Figure S4. The morphology of the suspended cell spheres formed in serum-free neural stem cell medium composed of DMEM/F12, 20 ng/ml EGF, 20 ng/ml bFGF, and 1x B27. Figure S5. The levels of the indicated cytokines were assessed by ELISA. T cells were incubated with GSC-3# cells at an E:T ratio of 5:1. The results are presented as the mean volume ± SD, ***, P < 0.001; ns, not significant. Figure S6. NKG2D-BBz CAR-T cells lysed U-87MG cells effectively in mice. (A) B-NDG mice were injected with 1 × 106 stable luciferase transfected U-87MG cells subcutaneously and imaged 7 days prior to T cell infusion. After mice received T cells treatment, photographs were taken serially at indicated time. (B) Comparison of tumor bioluminescent signal among the indicated groups at different time points. Figure S7. Persistence of NKG2D-BBz CAR-T cells in mice. B-NDG mice were injected with 1 × 106 stable luciferase transfected U-87MG cells subcutaneously and received T cells treatment 7 days later. Then human genomic DNA in blood was detected using qPCR at indicated time. Figure S8. Growth curves for the indicated cells. The CAR-T cells were counted every 2 days. The data are presented as the mean ± SD; ns, not significant. (DOCX 3450 kb)