Kayadiol exerted anticancer effects through p53-mediated ferroptosis in NKTCL cells.

BACKGROUND: Extranodal natural killer/T cell lymphoma (NKTCL) is a highly aggressive type of non-Hodgkin lymphoma that facing the treatment challenges. Natural compounds are important sources for drug development because of their diverse biological and chemical properties, among which terpenoids have strong anticancer activities. METHODS: The human NK/T cell lymphoma cell line YT and peripheral blood lymphocytes isolated from NKTCL patients were treated with different concentrations of kayadiol. Then, the following experiments were performed: CCK-8 assay for cell viability, reactive oxygen species (ROS) and glutathione (GSH) assay and co-treatment with NAC, reduced GSH, or ferrostatin-1 for ferroptosis, the proteome profiling for elucidating signaling pathways, and western blot for the expression of p53, SCL7A11, and GPX4. siRNA and CRISPR/Cas9 plasmid for p53 knockout was designed and transfected into YT cells to evaluate the causal role of p53 in kayadiol-induced ferroptosis. The synergistic effect was evaluated by CCK8 assay after co-treatment of kayadiol with L-asparaginase or cisplatin. RESULTS: In this study, we found that kayadiol, a diterpenoid extracted from Torreya nucifera, exerted significant killing effect on NKTCL cells without killing the healthy lymphocytes. Subsequently, we observed that kayadiol treatment triggered significant ferroptosis events, including ROS accumulation and GSH depletion. ROS scavenger NAC, GSH, and ferroptosis inhibitor ferrostatin-1 (Fer-1) reversed kayadiol-induced cell death in NKTCL cells. Furthermore, kayadiol decreased the expression of SLC7A11 and GPX4, the negative regulatory proteins for ferroptosis. We then demonstrated that p53 was the key mediator of kayadiol-induced ferroptosis by SLC7A11/GPX4 axis through p53 knockout experiments. In addition, kayadiol exerted a synergistic effect with L-asparaginase and cisplatin in NKTCL cells. CONCLUSION: Taken together, our results suggested that the natural product kayadiol exerted anticancer effects through p53-mediated ferroptosis in NK/T cell lymphoma cells. Hence, it can serve as an effective alternative in the treatment of NK/T cell lymphoma, especially for patients exhibiting chemoresistance.

Inhibition of SRPK1, a key splicing regulator, exhibits antitumor and chemotherapeutic-sensitizing effects on extranodal NK/T-cell lymphoma cells.

BACKGROUND: Increasing evidence has convincingly shown that abnormal pre-mRNA splicing is implicated in the development of most human malignancies. Serine/arginine-rich protein kinase 1 (SRPK1), a key splicing regulator, is reported to be overexpressed in leukemia and other cancer types, which suggests the therapeutic potential of targeting SRPK1. METHODS: SRPK1 expression was measured in 41 ENKTL patients by immunohistochemistry and mRNA expression was analyzed by qRT‒PCR. We knocked down SRPK1 expression in the ENKTL cell line YT by siRNA transfection and inhibited SRPK1 using inhibitors (SPHINX31 and SRPIN340) in YT cells and peripheral blood lymphocytes (PBLs) isolated from ENKTL patients to investigate its role in cell proliferation and apoptosis. Then, RNA-seq analysis was performed to predict the potential signaling pathway by which SRPK1 inhibition induces cell death and further verified this prediction by Western blotting. RESULTS: In the present study, we initially evaluated the clinical significance of SRPK1 in extranodal natural killer/T-cell lymphoma (ENKTL), a very aggressive subtype of non-Hodgkin lymphoma. The expression of SRPK1 in ENKLT patients was examined by immunohistochemistry and qRT‒PCR, which revealed SRPK1 overexpression in more than 60% of ENKTL specimens and its association with worse survival. Cellular experiments using the human ENKTL cell line YT and PBLs from ENKTL patients, demonstrated that inhibition of SRPK1 suppressed cell proliferation and induced apoptosis. Subsequently, we investigated the downstream targets of SRPK1 by RNA-seq analysis and found that SRPK1 inhibition induced ATF4/CHOP pathway activation and AKT1 inhibition. Furthermore, ENKTL patients presenting high SRPK1 expression showed resistance to cisplatin-based chemotherapy. The association of SRPK1 expression with cisplatin resistance was also confirmed in YT cells. SRPK1 overexpression via pLVX-SRPK1 plasmid transfection dramatically decreased the sensitivity of YT cells to cisplatin, while siRNA-mediated SRPK1 knockdown or SRPK1 inhibitor treatment significantly increased cisplatin cytotoxicity. CONCLUSION: In summary, these results support that SRPK1 might be a useful clinical prognostic indicator and therapeutic target for ENKTL, especially for patients who relapse after cisplatin-based chemotherapies.

Clinical characteristics of 3062 COVID-19 patients: A meta-analysis.

We aimed to systematically review the clinical characteristics of coronavirus disease 2019 (COVID-19). Seven databases were searched to collect studies about the clinical characteristics of COVID-19 from January 1, 2020 to February 28, 2020. Then, meta-analysis was performed by using Stata12.0 software. A total of 38 studies involving 3062 COVID-19 patients were included. Meta-analysis showed that a higher proportion of infected patients was male (56.9%). The incidence rate of respiratory failure or acute respiratory distress syndrome was 19.5% and the fatality rate was 5.5%. Fever (80.4%), fatigue (46%), cough (63.1%), and expectoration (41.8%) were the most common clinical manifestations. Other common symptoms included muscle soreness (33%), anorexia (38.8%), chest tightness (35.7%), shortness of breath (35%), dyspnea (33.9%). Minor symptoms included nausea and vomiting (10.2%), diarrhea (12.9%), headache (15.4%), pharyngalgia (13.1%), shivering (10.9%), and abdominal pain (4.4%). The proportion of patients that was asymptomatic was 11.9%. Normal leukocyte counts (69.7%), lymphopenia (56.5%), elevated C-reactive protein levels (73.6%), elevated ESR (65.6%), and oxygenation index decreased (63.6%) were observed in most patients. About 37.2% of patients were found with elevated D-dimer, 25.9% of patients with leukopenia, along with abnormal levels of liver function (29%), and renal function (25.5%). Other findings included leukocytosis (12.6%) and elevated procalcitonin (17.5%). Only 25.8% of patients had lesions involving a single lung and 75.7% of patients had lesions involving bilateral lungs. The most commonly experienced symptoms of COVID-19 patients were fever, fatigue, cough, and expectoration. A relatively small percentage of patients were asymptomatic. Most patients showed normal leucocytes counts, lymphopenia, elevated levels of C-reactive protein and ESR. Bilateral lung involvement was common.

Nuclear factor of activated T cells as a marker of in vivo low-dose dibenzo[a,h]anthracene exposure.

We previously demonstrated that particulate matter ≤2.5 µm (PM2.5) suppresses the immune response in the spleen in vivo. Although PM2.5 includes the polycyclic aromatic hydrocarbon (PAH) such as dibenzo[a,h]anthracene (DBA), it is unclear whether PAH has a direct effect on the responses of splenocytes. In our study, the concentration of DBA used was approximately 0.8 µm, which is much lower than concentrations used in other toxicological studies of DBA. Although exposure to high concentrations of DBA is implicated in carcinogenesis, the effects of low doses of DBA on immune cells in vivo remain unclear. Here, we investigated the effects of low DBA doses on mouse splenocytes in vivo. Mice were administered dimethyl sulfoxide or DBA (0.4 or 0.8 µm) intratracheally. Twenty-four hours after treatment, the mice were killed and their splenocytes were collected. DBA treatment enhanced mitogen-induced cell proliferation and cytokine production in the mouse splenocytes. Furthermore, DBA enhanced splenic CD4+ and CD8+ cell proliferation and cytokine production. The nuclear factor of activated T cells (NFAT) was activated in CD4+ cells. DBA also activated nuclear factor-kappa B and CCAAT enhancer-binding protein pathways in CD11b+ cells. DBA-enhanced splenocyte activation was Toll-like receptor 2-, 4-, 9- and MyD88-independent. These results suggest that NFAT represents a promising marker for evaluation of the effects of DBA on T cells and T-cell-dependent antibody responses.

Lipopolysaccharide levels adherent to PM2.5 play an important role in particulate matter induced-immunosuppressive effects in mouse splenocytes.

Epidemiological studies show that exposure to ambient particulate matter (PM) is associated with serious adverse health effects, including, but not limited to, those on the respiratory system. In the present study, we investigated the splenic response in mice administered PM of ≤ 2.5 µ m diameter (PM2.5). Male BALB/c mice (7 or 8 weeks old) were intratracheally administered PM2.5 (0.1 mg) four times, at 2 week intervals, and dissected 24 h after the final administration. The effect of six types of PM2.5, collected in Shenyang or Beijing (China) and Kitakyushu (Japan), on splenocytes was examined. Our results revealed a strong correlation between the levels of lipopolysaccharide (LPS), but not that of ß-glucan and polycyclic aromatic hydrocarbons, attached to PM2.5 and the effect of PM2.5 on cell activity. PM2.5 with a low amount of LPS (PM2.5LL) reduced splenocyte mitogen-induced proliferation and cytokine production compared with that in control mice. The suppressive effects of PM2.5LL on proliferation and interleukin-2 production in splenocytes were rescued by the antioxidant N-acetylcysteine. Expression of heme oxygenase-1 was elevated after PM2.5LL administration, particularly in CD11b +  cells, while no elevation was observed in CD4+ , CD8+ or B220+ cells. Further, dissociation of the nuclear factor erythroid 2-related factor 2 from Kelch-like ECH-associating protein 1 was observed in splenocytes of PM2.5LL-administered mice. These data suggest that LPS attached to PM2.5 modulates the splenocyte immune responses to PM2.5.

PM2.5-induced lung inflammation in mice: Differences of inflammatory response in macrophages and type II alveolar cells.

Particulate matter 2.5 (

The Treatment of Diffuse Large B-Cell Lymphoma (Triple Expression) Involving the Breast, Spleen, and Bone in a Male Patient with Viral Hepatitis B: A Rare Case Report.

Background: Diffuse large B-cell lymphoma (DLBCL) involving the breast, spleen, and bone in a male patient with hepatitis B virus (HBV) infection is extremely rare in clinical practice. Case Presentation: We report a case of DLBCL involving the breast, spleen, and bone (triple expression of Bcl-2+, Bcl-6+, and 70% positive C-mcy) in a male patient with HBV admitted to our hospital. The patient was treated with EPOCH×4, lenalidomide+EPOCH×2 chemotherapy, intermittent methotrexate intrathecal injections to prevent central invasion, and autologous stem cell transplantation (ASCT). The patient is currently in complete remission, and the follow-up time was 43 months. Conclusion: A patient with DLBCL involving the breast, spleen, and bone can be treated with a combination of multiple regimens. If the patient's economic conditions permit it, ASCT can be considered.

Targeted sequencing reveals the relationship between mutations and patients' clinical indicators, blood cell counts and early progression in diffuse large-B cell lymphoma.

In the current study, we assessed the relationship between mutations and the blood cell counts and early progression of patients with diffuse large-B cell lymphoma (DLBCL). A total of 109 patients with newly diagnosed DLBCL were included in this study. UBE2A mutation was only found in patients with bone marrow involvement. The mutations of ZNF608, SF3B1, DTX1, and NCOR2 were related to blood cell counts. NCOR2 mutations were only detected in patients of the noncomplete response group (PR + SD + PD). In addition, the mutations of ATM, BTG2, TBL1XR1, and TP53 were linked to lower PFS/OS rate, while SGK1, SCOS1, and NFKBIE were related to higher PFS/OS rate. Importantly, we identified that Ann Arbor stage (III-IV), B symptoms, absolute lymphocyte count (ALC) abnormity, and MTOR mutation were the four independent influencing factors of the 12-month progression of DLBCL patients. Overall, this study revealed that mutations were associated with the early progression of DLBCL.

Association Between Polymorphism in Diabetes Susceptibility Gene Insulin-Like Growth Factor 2mRNA-Binding Protein 2 and Risk of Diffuse Large B-Cell Lymphoma.

Background: Numerous studies have shown that polymorphisms in the diabetes susceptibility gene, insulin-like growth factor 2mRNA-binding protein 2 (IGF2BP2), are associated with the occurrence and development of various malignant tumors; however, their correlation with the onset of diffuse large B-cell lymphoma (DLBCL) is still unknown. Therefore, this study aimed to explore whether IGF2BP2 polymorphisms increase the risk of developing DLBCL. Methods: This study included 295 DLBCL patients and 331 healthy individuals. Peripheral blood was collected, and polymerase chain reaction-ligase detection reaction (PCR-LDR) was used to detect IGF2BP2 gene polymorphisms. Logistic regression was used to assess the association between IGF2BP2 polymorphism and the risk of DLBCL, adjusted for age, sex, and body mass index (BMI). P < .05 indicated statistical significance. Results: The rs4402960 polymorphism in the IGF2BP2 gene was associated with the occurrence and development of DLBCL. After adjusting for age, sex, and BMI, GT (odd ratio [OR] = 1.54; 95% confidence interval [CI] = 1.08-2.19; P = .016), TT (OR = 2.00; 95% CI = 1.09-3.68; P = .026), and T genotype carrying (GT + TT) (OR = 1.62; 95% CI = 1.17-2.25; P = .004) significantly increased the risk of DLBCL. This study also found that the polymorphism rs1470579 was related to the development of DLBCL. After adjusting for age, sex, and BMI, AC (OR = 1.55; 95% CI = 1.11-2.17; P = .010), CC (OR = 2.18; 95% CI = 1.17-4.06; P = .014), and C genotype carrying (AC + CC) (OR = 1.64; 95% CI = 1.19-2.26; P = .002) significantly increased the risk of DLBCL. Conclusions: Our study found that polymorphism in the IGF2BP2 gene was associated with an increased risk of developing DLBCL.

Influence of Hyperglycemia on the Prognosis of Patients with Diffuse Large B-Cell Lymphoma.

Purpose: To explore the effects of primary and secondary hyperglycemia and the application of the hypoglycemic drug metformin on the prognosis of patients with diffuse large B-cell lymphoma (DLBCL). Methods: We performed a retrospective analysis of 1767 DLBCL patients.Cox regression method was used for analysis to evaluate the prognostic factors, and the Kaplan-Meier method was used to draw a survival curve to analyze the effect of hyperglycemia and the hypoglycemic drug metformin on the progression-free survival (PFS) and overall survival (OS) of DLBCL patients. Results: Our study showed that patients with hyperglycemia tend to have higher age (age>60 years), high body mass index (BMI)(≥24kg/m2), late Ann Arbor stage (III-IV), high international prognostic index (IPI) (3-5 score), high lactic dehydrogenase (LDH) level (>250U/L), bulky disease and comorbidity. Hyperglycemia affects the survival time of the DLBCL population (PFS: adjusted HR 1.41, 95% CI: 1.16-1.70, P <0.001, OS: adjusted HR 1.33, 95% CI:1.09-1.61, P=0.004).Compared with the non-hyperglycemia group, the secondary hyperglycemia increase affects the prognosis of the DLBCL population (P<0.001). Compared with the secondary hyperglycemia group, the primary hyperglycemia group has a poor prognosis (P<0.05). For patients with DLBCL and hyperglycemia (732 patients in total), the use of metformin can improve their PFS and OS (PFS: adjusted HR 0.69, 95% CI: 0.49-0.96, P=0.028, OS: adjusted HR 0.68, 95% CI: 0.49-0.95, P=0.024). Conclusion: Hyperglycemia and secondary hyperglycemia are related to the poor prognosis of DLBCL population.For patients with DLBCL combined with hyperglycemia, the application of metformin can improve survival rate.

Identification of Genes with Altered Methylation and Its Role in Early Diagnosis of Sepsis-Induced Acute Respiratory Distress Syndrome.

PURPOSE: Early diagnosis of sepsis-induced acute respiratory distress syndrome (ARDS) is critical for effective treatment. We aimed to identify early stage biomarkers. MATERIALS AND METHODS: Differentially expressed genes were identified in whole blood samples from patients with sepsis or ARDS based on the Gene Expression Omnibus (GEO) datasets GSE32707, GSE54514 and GSE10361. Functional enrichment analysis explored the biological characteristics of differentially expressed genes. Genes with high functional connectivity based on a protein-protein interaction network were marked as hub genes, which were validated using the GEO dataset GSE76293, and a gene set variation analysis index (GSVA) was assigned. Diagnostic and predictive ability of the hub genes were assessed by receiver operating characteristic (ROC) curve analysis. DNA methylation levels of hub genes were quantified using the GEO dataset GSE67530. RESULTS: Forty-one differentially expressed genes were shared between sepsis-specific and ARDS-specific datasets. MAP2K2 and IRF7 functional activity was highly connected in sepsis-induced ARDS. Hub genes included RETN, MVP, DEFA4, CTSG, AZU1, FMNL1, RBBP7, POLD4, RIN3, IRF7. ROC curve analysis of the hub gene GSVA index showed good diagnostic ability in sepsis or ARDS. Among genes related to sepsis-induced ARDS, 17 were differentially methylated. Principal component analysis and heatmaps indicated that gene methylation patterns differed significantly between ARDS patients and controls. CONCLUSION: We identified a genetic profile specific to early-stage sepsis-induced ARDS. The abnormal expression of these genes may be caused by hypomethylation, which may serve as a biomarker for early diagnosis of ARDS.

Two gene set variation indexes as potential diagnostic tool for sepsis.

Accurate diagnosis of sepsis remains challenging, new markers or combinations of markers are urgently needed. In the present study, we screened differentially expressed genes (DEGs) between sepsis and non-sepsis blood samples across three previously published gene expression data sets. Common upregulated and downregulated DEGs were ranked according to their average functional similarity. The ten genes (OLFM4, ORM1, CEP55, S100A12, S100P, LRG1, CEACAM8, MS4A4A, PLSCR1, and IL1R2) with the largest average functional similarity among the common upregulated genes and another ten genes (THEMIS, IL2RB, CD2, IL7R, CD3E, KLRB1, PVRIG, CCRR3, TGFBR3, and PLEKHA1) with the largest average functional similarity among the common downregulated genes were separately identified as the upregulated crucial gene set and the downregulated crucial gene set. Gene set variation analysis (GSVA) was used to obtain the GSVA index of each sample against the two crucial gene sets. Both the two crucial GSVA indexes may be robust markers for sepsis with high area under ROC curve. The diagnostic utility of the upregulated GSVA index was validated in another independent data set. Functional analyses revealed several sepsis-related pathways. In conclusion, we proposed two sepsis-related gene sets across multiple data sets and created two GSVA indexes with promising diagnostic value.

Lipopolysaccharide attached to urban particulate matter 10 suppresses immune responses in splenocytes while particulate matter itself activates NF-κB.

We previously reported that Asian sand dust (ASD), which contains particulate matter (PM) less than 10 µm in diameter (PM10), induced subacute inflammation in splenocytes. However, it was unclear whether the PM itself or compounds attached to its surface induced the inflammation. Here we characterized the role of organic substances adsorbed onto the PM10 surface in triggering inflammation by comparing the effect on splenocyte activation of PM10 from urban areas (urPM10), which is rich in lipopolysaccharide (LPS) as compared to ASD, with that of heated PM10 (H-PM). BALB/c mice were intratracheally administered urPM10 or H-PM with or without LPS (1 ng and 10 ng) four times at 2-week intervals, and splenocytes were prepared at 24 h after the final administration to assay the immune responses. urPM10 suppressed splenocyte activation, while H-PM activated splenocytes and LPS neutralization by polymyxin B rescued urPM10-induced immunosuppression. Co-administration of LPS with H-PM clearly suppressed mitogen-induced immune responses in the spleen. Consistent with these results, H-PM induced the phosphorylation of nuclear factor κB (NF-κB) p65 and I kappa B kinase (IKK), which was inhibited by co-administration of LPS. In mice deficient in the LPS signal transducer MyD88, splenocyte activation after LPS or H-PM treatment in vivo was comparable to that in the control. Altogether, our results indicate that PM10 itself could activate NF-κB through the MyD88 pathway, which was modulated by the amount of LPS attached.

Autophagy is associated with cucurbitacin D-induced apoptosis in human T cell leukemia cells.

We previously reported that the inflammasome inhibitor cucurbitacin D (CuD) induces apoptosis in human leukemia cell lines. In the present study, we investigated the effects of co-treatment with an additional Bcl-xL inhibitor, Z36. Treatment with Z36 induced cell death in leukemia cell lines, with MT-4 cells exhibiting the lowest sensitivity to Z36. Co-treatment of cells with Z36 and CuD resulted in a greater degree of cell death for Hut78 and Jurkat cells than treatment with CuD alone. In contrast, co-treatment of MT-4 cells with Z36 and CuD had a suppressive effect on cell death. The autophagy inhibitor 3-methyladenine (3-MA) suppressed the growth of leukemia cell lines HuT78, Jurkat, MT-1, and MT-4. CuD-induced cell death was enhanced by 3-MA in Jurkat cells, but inhibited in MT-4 cells. Western blotting results revealed cleavage of poly(ADP ribose) polymerase (PARP), supporting CuD-induced cell death; 3-MA enhanced CuD-Z36-induced PARP cleavage. Taken together, our results indicate that autophagy negatively regulates chemical-induced cell death of leukemia cells, and that controlling autophagy could be beneficial in the development of more effective chemotherapies against leukemia.

Relationship between triterpenoid anticancer drug resistance, autophagy, and caspase-1 in adult T-cell leukemia.

We previously reported that the inflammasome inhibitor cucurbitacin D (CuD) induces apoptosis in human leukemia cell lines. Here, we investigated the effects of CuD and a B-cell lymphoma extra-large (Bcl-xL) inhibitor on autophagy in peripheral blood lymphocytes (PBL) isolated from adult T-cell leukemia (ATL) patients. CuD induced PBL cell death in patients but not in healthy donors. This effect was not significantly inhibited by treatment with rapamycin or 3-methyladenine (3-MA). The Bcl-xL inhibitor Z36 induced death in primary cells from ATL patients including that induced by CuD treatment, effects that were partly inhibited by 3-MA. Similarly, cell death induced by the steroid prednisolone was enhanced in the presence of Z36. A western blot analysis revealed that Z36 also promoted CuD-induced poly(ADP ribose) polymerase cleavage. Interestingly, the effects of CuD and Z36 were attenuated in primary ATL patient cells obtained upon recurrence after umbilical cord blood transplantation, as compared to those obtained before chemotherapy. Furthermore, cells from this patient expressed a high level of caspase-1, and treatment with caspase-1 inhibitor-enhanced CuD-induced cell death. Taken together, these results suggest that rescue from resistance to steroid drugs can enhance chemotherapy, and that caspase-1 is a good marker for drug resistance in ATL patients.

PM2.5 collected in China causes inflammatory and oxidative stress responses in macrophages through the multiple pathways.

Air pollution continues to increase in East Asia, particularly in China, and is considered to cause serious health problems. In this study, we investigated the toxicological properties of particulate matter ≤2.5mm (PM2.5) collected in an urban area in China (Shenyang), focusing on inflammation and oxidative stress tightly linked to respiratory diseases. Exposure to PM2.5 significantly increased the expression levels of inflammatory (interleukin-1ß and cyclooxygenase-2) and oxidative stress (heme oxygenase1) genes in the mouse macrophages. PM2.5-caused inflammatory response was strongly suppressed by endotoxin neutralizer (polymyxin B) and knock-out of toll-like receptor 4, while oxidative stress was not. On the other hand, an antioxidant (N-acetylcystein) suppressed oxidative stress, but not inflammatory response. These results suggest that PM2.5 in the atmospheric environment of China causes inflammation and oxidative stress in macrophages via separate pathways.