Depletion of Gr1+ myeloid cells attenuates high-fat-diet-aggravated esophageal squamous cell carcinoma in mice.

Obesity is a leading cause of multiple cancers, but whether it promotes esophageal squamous cell carcinoma (ESCC) and its underlying cancer-promoting mechanism remains unclear. To evaluate the effect of a high-fat diet (HFD) on ESCC and explore the role and mechanism of myeloid-derived suppressor cells (MDSCs) in HFD-induced ESCC, C57BL/6J mice were treated with 4-nitroquinoline 1-oxide (4NQO) to induce ESCC and randomly assigned to an HFD or a normal-fat diet. An anti-Gr1 monoclonal antibody was used to deplete MDSCs in the context of experimental diets and ESCC induction. The expression of MDSC markers CD11b and Gr1 and immune checkpoints (ICs) PD1, TIM3, and VISTA in lesions were detected by immunohistochemistry. The correlation between myeloid cell markers (CD11b and CD33) and ICs and their relationship with ESCC patient prognosis were further analyzed using the The Cancer Genome Atlas dataset. The results showed that HFD accelerated esophageal carcinogenesis, induced MDSC expansion, and upregulated IC expression, whereas depletion of Gr1+ myeloid cells significantly suppressed tumor growth, decreased the number of MDSCs, and downregulated IC expression in HFD mice. PD1, TIM3, and VISTA expressions were positively correlated with myeloid cell marker expression in human ESCC. Moreover, the high expression of IC molecules was associated with poor survival in patients with ESCC. These data indicate that HFD promotes the initiation and development of ESCC. Gr1+ myeloid cell targeting significantly inhibited ESCC formation in HFD mice, which may be associated with IC downregulation.

Lipopolysaccharide inhibits GPR120 expression in macrophages via Toll-like receptor 4 and p38 MAPK activation.

Free fatty acid receptor G protein-coupled receptor 120 (GPR120) is highly expressed in macrophages and was reported to inhibit lipopolysaccharide (LPS)-stimulated cytokine expression. Under inflammation, macrophages exhibit striking functional changes, but changes in GPR120 expression and signaling are not known. In this study, the effects of LPS treatment on macrophage GPR120 expression and activation were investigated. The results showed that LPS inhibited GPR120 expression in mouse macrophage cell line Ana-1 cells. Moreover, LPS treatment inhibited GPR120 expression in mouse alveolar macrophages both in vitro and in vivo. The inhibitory effect of LPS on GPR120 expression was blocked by Toll-like receptor 4 (TLR4) inhibitor TAK242 and p38 mitogen-activated protein kinase inhibitor LY222820, but not by ERK1/2 inhibitor U0126 and c-Jun N-terminal kinase inhibitor SP600125. LPS-induced inhibition of GPR120 expression was not attenuated by GPR120 agonists TUG891 and GW9508. TUG891 inhibited the phagocytosis of alveolar macrophages, and LPS treatment counteracted the effects of TUG891 on phagocytosis. These results indicate that pretreatment with LPS inhibits GPR120 expression and activation in macrophages. It is suggested that LPS-induced inhibition of GPR120 expression is a reaction enhancing the LPS-induced pro-inflammatory response of macrophages.

Interplay of Toll-Like Receptor 9, Myeloid Cells, and Deubiquitinase A20 in Periodontal Inflammation.

Toll-like receptor 9 (TLR9)-deficient (TLR9-/-) mice are resistant to periodontitis, a disease characterized by a dysbiotic microbiota and deregulated immune response and resulting in tooth loss and various systemic conditions. However, the mechanisms and biological pathways by which TLR9 instigates periodontal inflammation are yet to be identified. In a ligature-induced model of periodontitis, we demonstrate that TLR9-/- mice exhibited significantly less alveolar bone loss than their wild-type (WT) counterparts. Consistent with the disease phenotype, gingival tissues showed significantly more inflammatory cell infiltration in the WT ligated but not in the TLR9-/- ligated mice compared to the unligated controls. The peritoneal infection model using Porphyromonas gingivalis, a keystone pathogen for periodontitis, revealed reduced neutrophils in TLR9-/- mice on day 1 postinfection compared to the levels in WT mice. Transcriptomics analyses showed increased expression of A20 (tumor necrosis factor alpha [TNF-α]-induced protein 3 [TNFAIP3]), an inhibitor of the NF-κB pathway and a negative regulator of TLR signaling, in ligated TLR9-/- mouse gingival tissues compared to its expression in the WT. Ex vivo, TLR9-/- bone marrow-derived macrophages produced more A20 than WT cells following P. gingivalis challenge. Clinically, A20 was modestly upregulated in human gingival tissue specimens from chronic periodontitis patients, further confirming the biological relevance of A20 in periodontal inflammation. We conclude that TLR9 modulates periodontal disease progression at both the cellular and molecular level and identify A20 as a novel downstream signaling molecule in the course of periodontal inflammation. Understanding the regulation of the TLR9 signaling pathway and the involvement of A20 as a limiting factor of inflammation will uncover alternative therapeutic targets to treat periodontitis and other chronic inflammatory diseases.

Reactive oxygen species mediated T lymphocyte abnormalities in an iron-overloaded mouse model and iron-overloaded patients with myelodysplastic syndromes.

The adverse effects of iron overload have raised more concerns as a growing number of studies reported its association with immune disorders. This study aimed to investigate alterations in the immune system by iron overload in patients with myelodysplastic syndrome (MDS) and an iron-overloaded mouse model. The peripheral blood from patients was harvested to test the effect of iron overload on the subsets of T lymphocytes, and the level of reactive oxygen species (ROS) was also evaluated. The data showed that iron-overloaded patients had a lower percentage of CD3+ T cells and disrupted T cell subsets, concomitant with higher ROS level in lymphocytes. In order to explore the mechanism, male C57Bl/6 mice were intraperitoneally injected with iron dextran at a dose of 250 mg/kg every 3 days for 4 weeks to establish an iron-overloaded mouse model and the blood of each mouse was collected for the analysis of the T lymphocyte subsets and T cell apoptosis. The results showed that iron overload could reduce the percentage of CD3+ T cells and the ratio of Th1/Th2 and Tc1/Tc2 but increase the percentage of regulatory T (Treg) cells and the ratio of CD4/CD8. We also found that iron overload induced the apoptosis of T lymphocytes and increased its ROS level. Furthermore, these effects could be partially recovered after treating with antioxidant N-acetyl-L-cysteine (NAC) or iron chelator deferasirox (DFX). Taken together, these observations indicated that iron overload could selectively affect peripheral T lymphocytes and induce an impaired cellular immunity by increasing ROS level.

[Capsaicin Alleviate Myocardial Ischemia Reperfusion Injury Through Attenuating Mitochondrial Oxidative Stress].

OBJECTIVE: To investigate the role of capsaicin (CAP) in myocardial ischemia reperfusion injury and its underlying mechanisms. METHODS: Twentyfour adult male SD rats were randomized into 4 groups,namely the control group,ischemia reperfusion group,ischemia reperfusion with CAP group,and ischemia reperfusion with CAPZ and CAP group. Isolated rat hearts underwent Langendorff perfusion. Left ventricular enddiastolic pressure (LVEDP) andleft ventricular developed pressure (LVDP) was calculated to evaluate myocardial performance at 30 min of reperfusion.Triphenyltetrazolium chloride staining was used to measure the infarct size of myocardium at 120 min reperfusion. The morphological changes in myocardial fiber was analyzed by HE staining at the end of reperfusion. Lactate dehydrogenase (LDH) content in the coronary flow was determined during the first 5 min reperfusion. The myocardial mitochondria was isolated and extracted for measuring a series of indicators of mitochondrial oxidative stress,including superoxide dismutase (SOD),methane dicarboxylic aldehyde (MDA) at the end of reperfusion. Western blot was used to determine the expression of caspase3 and cytochrome c at 10 min reperfusion. RESULTS: Compared with the control group,IR group significantly decreased in cardiac function,the level of LVDP and SOD activity and induced an enlarged infarct size ( P<0.01),accompanied by the disordered arrangement of myocardial cells,the content of MDA was increased ( P<0.01),the content of caspase3 and cytochrome c were also upregulated ( P<0.01).10 µmol/L CAP significantly attenuated these effects induced by ischemia reperfusion injury,levels of LVDP and infarct size at the end of reperfusion were significantly improved( P<0.01),nevertheless levels of LVEDP and MDA at the end of reperfusion and LDH were down-regulated markedly ( P<0.01),the content of caspase-3 and cytochrome c were also decreased ( P<0.01). CONCLUSION: These results demonstrate that CAP can suppresses cell apoptosis and necrosis,and alleviate heart function and cell survival from ischemia reperfusion injury through attenuating mitochondrial oxidative stress.

Toll-Like Receptor 9-Mediated Inflammation Triggers Alveolar Bone Loss in Experimental Murine Periodontitis.

Chronic periodontitis is a local inflammatory disease induced by a dysbiotic microbiota and leading to destruction of the tooth-supporting structures. Microbial nucleic acids are abundantly present in the periodontium, derived through release after phagocytic uptake of microbes and/or from biofilm-associated extracellular DNA. Binding of microbial DNA to its cognate receptors, such as Toll-like receptor 9 (TLR9), can trigger inflammation. In this study, we utilized TLR9 knockout (TLR9(-/-)) mice and wild-type (WT) controls in a murine model of Porphyromonas gingivalis-induced periodontitis and report the first in vivo evidence that TLR9 signaling mediates the induction of periodontal bone loss. P. gingivalis-infected WT mice exhibited significantly increased bone loss compared to that in sham-infected WT mice or P. gingivalis-infected TLR9(-/-) mice, which were resistant to bone loss. Consistent with this, the expression levels of interleukin 6 (IL-6), tumor necrosis factor (TNF), and receptor-activator of nuclear factor kappa B ligand (RANKL) were significantly elevated in the gingival tissues of the infected WT mice but not in infected TLR9(-/-) mice compared to their levels in controls. Ex vivo studies using splenocytes and bone marrow-derived macrophages revealed significantly diminished cytokine production in TLR9(-/-) cells relative to the cytokine production in WT cells in response to P. gingivalis, thereby implicating TLR9 in inflammatory responses to this organism. Intriguingly, compared to the cytokine production in WT cells, TLR9(-/-) cells exhibited significantly decreased proinflammatory cytokine production upon challenge with lipopolysaccharide (LPS) (TLR4 agonist) or Pam3Cys (TLR2 agonist), suggesting possible cross talk between TLR9, TLR4, and TLR2. Collectively, our results provide the first proof-of-concept evidence implicating TLR9-triggered inflammation in periodontal disease pathogenesis, thereby identifying a new potential therapeutic target to control periodontal inflammation.

Antiviral butyrolactones from the endophytic fungus Aspergillus versicolor.

Versicolactones A-D (1-4), four new butyrolactones, along with four known butyrolactones (5-8) were isolated from the fermentation products of the endophytic fungus Aspergillus versicolor. The structures of compounds 1-4, including absolute configuration, were elucidated by interpretation of the NMR and CD data. Compound 2 was further confirmed by single-crystal X-ray diffraction analysis. In particular, compound 1 is the first naturally occurring butyrolactone possessing an unusual 2-oxopropyl group. More importantly, compounds 1 and 8 displayed significant antitobacco mosaic virus activities with inhibition rates of 46.4 % and 35.4 %, even more potent than the positive control ningnanmycin (30.8 %). Compound 1 also showed moderate cytotoxicity against A549 and MCF7 cells with IC50 values of 3.2 and 2.5 µM, respectively.

7,8-Secolignans from the fruits of Schisandra neglecta.

Two new 7,8-secolignans, neglectahenols E and F (1 and 2), together with four known 7,8-secolignans (3-6), were isolated from the fruits of Schisandra neglecta. The structures were elucidated by spectroscopic methods, including extensive 1D and 2D NMR techniques. Compounds 1-6 were tested for their anti-tobacco mosaic virus (anti-TMV) activities at the concentration of 20 µM. Compounds 1 and 6 showed high anti-TMV activities with inhibition rates of 38.2% and 32.7%, respectively. These rates are higher than that of a positive control. Compounds 2-5 also showed modest anti-TMV activities with inhibition rates in the range of 22.8-28.7%. These rates are close to that of a positive control.

A retrospective cohort study on the protective effect of influenza vaccination during pregnancy in Gansu Province.

To evaluate the Vaccine Effectiveness (VE) of the Influenza Vaccine (InfV) during pregnancy in Gansu province. A retrospective cohort of pregnant women vaccinated with InfV was constructed using the electronic case system and immunization planning system of Gansu province. Outcome information of Influenza-Like Illness (ILI) in pregnant women after vaccination with InfV during the influenza epidemic seasons from 2019 to 2021 was collected. The log-binomial model was used to estimate the adjusted Risk Ratio (RR), VE, and their Confidence Intervals (CI). The incidence rates of the InfV vaccination group and the non-vaccination group during pregnancy in Gansu province were 4.70% (54/1,150) and 6.91% (16,990/245,754), respectively. The adjusted RR was 0.78 (95%CI 0.43-0.75), and the adjusted VE was 22.20 (95%CI 0.79-40.30). Vaccination with InfV during pregnancy in Gansu province from 2019 to 2021 has a protective effect against ILI in pregnant women, and there is no difference in vaccination at different stages of pregnancy.

Lateral reticular nucleus modulates the cardiosomatic reflex evoked by intrapericardial capsaicin in the rat.

The current study examined the role of the lateral reticular nucleus (LRN) in modulating the cardiosomatic reflex (CSR) induced by intrapericardial capsaicin in the anesthetized rat. Intrapericardial capsaicin was administered, and the CSR was monitored via electromyogram responses of the dorsal spinotrapezius muscle. Electrical stimulation of the LRN (10, 20 and 30 µA) depressed the CSR induced by intrapericardial capsaicin in an intensity-dependent manner. Microinjection of glutamate (4, 10, 20 and 40 nmol, in 0.2 µL) into the LRN replicated the effects of electrical stimulation. Furthermore, bilateral transections of the dorsolateral funiculus (DLF) decreased the LRN electrical stimulation-induced inhibition of the electromyogram responses. Intrathecal administration of the α2 -adrenergic receptor antagonist yohimbine or the serotonergic receptor antagonist methysergide significantly attenuated the LRN electrical stimulation-induced inhibition of the electromyogram responses. However, intrathecal application of the opioid receptor antagonist naloxone had no effect on the LRN electrical stimulation-induced inhibition. These results suggest that the LRN-DLF-spinal cord pathway is involved in descending inhibition of the CSR, and spinal α2 -adrenergic and serotonergic receptors participate in this descending inhibition.

MK-2206 induces cell cycle arrest and apoptosis in HepG2 cells and sensitizes TRAIL-mediated cell death.

It has become evident that AKT inhibitors have great potential in cancer treatment. In this study, we investigate the anticancer activity of MK-2206, a novel AKT inhibitor, on HepG2 hepatocellular carcinoma cell, and to show whether MK-2206 enhances the apoptosis-inducing potential of tumor necrosis factor-related apoptosis-inducing ligand (TRAIL). The cell growth inhibition was evaluated by MTT assay and colony formation assay. Cell cycle distribution was assessed by propidium iodide flow cytometry. Apoptosis was determined by AnnexinV-FITC/PI double staining assay and caspase-9, casapse-7, caspase-3, and PARP cleavage. The results of present study showed that MK-2206-induced G1-phase arrest was associated with a marked decrease in the protein expression of cyclin D1 with concomitant induction of p21 and p27. MK-2206-induced apoptosis was characterized by cleavage of a pro-caspase in a concentration-dependent manner. Moreover, the MAP family kinases p38 kinase and JNK were activated by exposure to MK-2206. SB203580, an p38-specific inhibitor, partially blocked MK-2206-induced death of HepG2 cells and caspase activation. A combination of MK-2206 with TRAIL significantly inhibited growth of TRAIL resistant HepG2 cells. Taken together, our findings provide a new insight to better understand anticancer mechanisms of MK-2206, at least in HepG2 cell. Using of MK-2206 as a potent sensitizer to TRAIL-induced apoptotic cell death offers a promising means of enhancing the efficacy of TRAIL-based HCC treatments.

[Reactive oxygen species mediate the injury and deficient hematopoietic supportive capacity of umbilical cord derived mesenchymal stem cells induced by iron overload].

OBJECTIVE: To explore the effects of iron overload on umbilical cord derived mesenchymal stem cells (UC-MSC) and elucidate the involvement of reactive oxygen species (ROS) in this process. METHODS: The iron overload model of MSC was established by in vitro addition of ferric ammonium citrate (FAC) into culture medium. Cell proliferation and apoptosis were determined by Annexin V/PI double staining and population doubling time (DT) respectively. Co-culture system was used to assess the hematopoietic support capacity of UC-MSC in different groups. Thereafter the ROS level was detected with fluorescent probe 2', 7'-dichlorofluorescin diacetate (DCFH-DA). And the ROS related signaling factors of p-p38MAPK, p38 MAPK, P53 were measured by Western blot. RESULTS: (1) The DT of UC-MSC in iron overload group was significantly longer than that of control ((24.43 ± 2.72) h vs (16.03 ± 2.31) h, P < 0.05). But the difference was insignificant after two passages (P > 0.05). (2) Apoptosis in iron overload group was higher than that of control (12.75% ± 0.32% vs 3.63% ± 0.80%, P < 0.05). (3) The colony forming capacity of mononuclear cell (MNC) co-cultured with UC-MSC of iron overload group for 1/2 weeks significantly decreased. (4) The ROS level of UC-MSC with iron overload was higher than that of control in time and concentration-dependent fashions and it peaked at 400 µmol/L of FAC for 12 h (1499 ± 86 vs 548 ± 97, P < 0.05). (5) The expressions of p-p38MAPK and P53 increased in response to FAC compared with control. But such an effect was partially inhibited after the use of antioxidants. CONCLUSIONS: Iron overload may impair the proliferation, survival and hematopoiesis supportive function of UC-MSC by enhancing the generation of ROS. And ROS stimulates the signaling pathways of p-p38MAPK and P53.

[Establishment of an mouse model of iron-overload and its impact on bone marrow hematopoiesis].

OBJECTIVE: To establish a mouse model of iron overload by intraperitoneal injection of iron dextran and investigate the impact of iron overload on bone marrow hematopoiesis. METHODS: A total of 40 C57BL/6 mice were divided into control group, low-dose iron group (12.5 mg/ml), middle-dose iron group (25 mg/ml), and high-dose iron group (50 mg/ml). The control group received normal saline (0.2 ml), and the rest were injected with intraperitoneal iron dextran every three days for six weeks. Iron overload was confirmed by observing the bone marrow, hepatic, and splenic iron deposits and the bone marrow labile iron pool. In addition, peripheral blood and bone marrow mononuclear cells were counted and the hematopoietic function was assessed. RESULTS: Iron deposits in bone marrow, liver, and spleen were markedly increased in the mouse models. Bone marrow iron was deposited mostly within the matrix with no significant difference in expression of labile iron pool.Compared with control group, the ability of hematopoietic colony-forming in three interventional groups were decreased significantly (P<0.05). Bone marrow mononuclear cells counts showed no significant difference. The amounts of peripheral blood cells (white blood cells, red blood cells, platelets, and hemoglobin) in different iron groups showed no significant difference among these groups;although the platelets were decreased slightly in low-dose iron group [(780.7±39.60)×10(9)/L], middle dose iron group [(676.2±21.43)×10(9)/L], and high-dose iron group [(587.3±19.67)×10(9)/L] when compared with the control group [(926.0±28.23)×10(9)/L], there was no significant difference(P>0.05). CONCLUSIONS: The iron-overloaded mouse model was successfully established by intraperitoneal administration of iron dextran. Iron overload can damage the hepatic, splenic, and bone marrow hematopoietic function, although no significant difference was observed in peripheral blood count.

[The role of NMDA and NK1 receptors in cardiac nociceptive information transmissions in the spinal cords of rats].

OBJECTIVE: To determine the effects of NMDA and NK1 receptor agonist and antagonist on the EMG and the synaptic mechanism of nociceptive information transmissions in the spinal cords. METHODS: Male SD rats were randomly divided into seven groups, with intrathecal injection of the following chemicals respectively: control group (10 microL saline), NMDA group (0.147 microg/10 pL NMDA), MK801 group (6.8 microg/10 microL MK801), MK801+NMDA group (6.8 microg/10 pL MK801+0. 147 microg/10 pL NMDA), Sar-SP group (1.4 pg/10 microL Sar-SP), CP-96345 group (5 microg/10 pL CP-96345), and CP-96345+Sar-SP group (1.4 micro/10 microL Sar-SP+5 microg/10 microL CP-96345). A cardiac pain model in rats through intrapericardial injection of capsaicin was established. Intrapericardial injection of capsaicin was given to the rats 10 min after intrathecal injection of the tested chemicals. The spinotrapezius electromyography (EMG) activities as an index of cardiac-somatic motor reflex were recorded simultaneously. RESULTS: Compared with the pre-test controls (100%), saline did not make a significant change to the capsaicin-evoked EMG response (96. 9% +/- 12. 5%, P>0. 05); NMDA agonist increased the capsaicin-evoked EMG response (185. 2% +/- 24. 4%) significantly (P<0. 05); neither MK801 nor a combined administration of MK801 and NMDA made a significant change to the capsaicin-evoked EMG response (106. 6% +/- 10. 2%, P> 0.05); Sar-SP increased the capsaicin-evoked EMG response (145. 6% 10. 1%) significantly (P<0. 05); whereas neither CP-96345 nor a combined administration of CP-96345 and Sar-SP made a significant change to the capsaicin-evoked EMG response (102. 2% +/- 8. 4%, P>0.05). CONCLUSION: NMDA and NK1 receptors may have participated in the transmissions of cardiac nociception information in the spinal cords of rats.

Effect of proton pump inhibitors on the risk of chronic kidney disease: A propensity score-based overlap weight analysis using the United Kingdom Biobank.

Background: Proton pump inhibitors (PPIs) are widely used and have been linked to kidney diseases. However, the role of PPI use in the development of chronic kidney disease (CKD) remains unclear. We undertook this study to examine the association between PPI use and the subsequent risk of CKD. Methods: This is a prospective analysis of 462,421 participants free of cancer diagnosis or chronic kidney disease from the United Kingdom Biobank. Self-reported PPI use was recorded using an electronic questionnaire and confirmed by a trained staff. Incident CKD was identified based on the medical history. Overlap propensity score weighting with the Cox model was used to calculate the effect of PPI use on CKD risk. The number needed to harm (NNH) was calculated at 5 and 10 years of follow-up. Results: We documented 7,031 cases of CKD over a median follow-up of 8.1 years. Overlap propensity score weighting analysis showed that regular PPI users had a 37% higher risk of CKD incident than non-users (HR 1.37, 95% CI 1.28-1.47). The association persisted across subgroup analyses, different types of PPIs, and several sensitivity analyses. Quantitative bias analysis indicated that the result was robust to unmeasured confounding (E-value 2.08, lower 95% CI 1.88). The NNH was 147.9 and 78.6 for 5 and 10 years of follow-up, respectively. A head-to-head comparison showed that PPI users had a 19% higher risk of CKD than H2RA users (HR 1.19, 95% CI 1.02-1.39). Conclusion: The regular use of PPI is associated with a higher risk of CKD. Healthcare providers should carefully weigh up the potential benefits against the risk in prescribing PPIs, particularly for patients requiring long-term treatment.

Evaluation of the safety and efficacy of humanized anti-CD19 chimeric antigen receptor T-cell therapy in older patients with relapsed/refractory diffuse large B-cell lymphoma based on the comprehensive geriatric assessment system.

Anti-CD19 chimeric antigen receptor (CAR) T-cell therapy has led to unprecedented results to date in relapsed/refractory (R/R) diffuse large B-cell lymphoma (DLBCL), yet its clinical application in elderly patients with R/R DLBCL remains somewhat limited. In this study, a total of 31 R/R DLBCL patients older than 65 years of age were enrolled and received humanized anti-CD19 CAR T-cell therapy. Patients were stratified into a fit, unfit, or frail group according to the comprehensive geriatric assessment (CGA). The fit group had a higher objective response (OR) rate (ORR) and complete response (CR) rate than that of the unfit/frail group, but there was no difference in the part response (PR) rate between the groups. The unfit/frail group was more likely to experience AEs than the fit group. The peak proportion of anti-CD19 CAR T-cells in the fit group was significantly higher than that of the unfit/frail group. The CGA can be used to effectively predict the treatment response, adverse events, and long-term survival.

[Effect and Involved Mechanism of RSL3-induced Ferroptosis in Acute Leukemia Cells MOLM13 and Drug-resistant Cell Lines].

OBJECTIVE: To investigate the effect and involved mechanism of RSL3 on ferroptosis action in acute leukemia cells MOLM13 and its drug-resistant cells. METHODS: After MOLM13 treated with RSL3, CCK-8 assay was performed to detect cell viability, flow cytometry was used to detect the reactive oxygen species (ROS) level of the cells, RT-qPCR and Western blot were used to detect the expression of glutathione peroxidase 4 (GPX4). After MOLM13/IDA and MOLM13/Ara-C, the drug-resistant cell lines were constructed, the ferroptosis induced by RSL3 was observed. Bone marrow samples were collected from patients with acute monocytic leukemia. RT-qPCR and Western blot were performed to detect the expression of related genes and proteins involved in ferroptosis pathway. RESULTS: RSL3 significantly inhibited the cell viability of MOLM13 and increased the intracellular ROS level, which were partially reversed by ferrostatin-1. The mRNA and protein expression of GPX4 decreased in MOLM13 treated with RSL3. RSL3 inhibited the viability of MOLM13/IDA and MOLM13/Ara-C cells more strongly than that of non-drug resistant cells, also increased the intracellular ROS level . The cytotoxic effects were partially reversed by ferrostatin-1. The mRNA and protein expressions of GPX4 in MOLM13/IDA and MOLM13/Ara-C cells were higher than those in non-drug resistant cells. The mRNA and protein levels of GPX4 in bone marrow of relapsed/refractory acute mononuclear leukemia patients were higher than those of ordinary acute mononuclear leukemia patients. CONCLUSION: RSL3 can induce non-drug resistant cells MOLM13 ferroptosis by inhibiting GPX4 activity. MOLM13/IDA and MOLM13/Ara-C are more sensitive to RSL3 compared with non-drug resistant cells MOLM13, which may be caused by the differences in GPX4 expression. The expressions of GPX4 mRNA and protein in relapsed/refractory acute mononuclear leukemia are higher than those in ordinary acute mononuclear leukemia.

Correlations of P21-activated kinase 1 expression to clinicopathological features of gastric carcinoma and patients' prognosis.

BACKGROUND AND OBJECTIVE: Recent studies proved that P21-activated kinase 1 (PAK1) is highly expressed in many kinds of tumor and plays an important role in genesis, development, and metastasis of tumor. We aimed to detect the expression of PAK1 in gastric carcinoma and to analyze its relationship with clinicopathological features and prognosis of gastric carcinoma. METHODS: Tissue microarray and immunohistochemical staining were performed to detect PAK1 in paraffin specimens of 189 gastric carcinomas, 54 paracancer tissues, 40 lymph nodes and 30 healthy tissues. Clinicopathologic features and follow-up data of the patients were analyzed by the Chi2 test and the Kaplan-Meier method. RESULTS: Positive rate of PAK1 was 73.0% in gastric carcinoma, 57.4% in paracancer tissues and 23.3% in healthy controls (Chi2 = 29.364, P < 0.05). Expression of PAK1 was significantly correlated with tumor size, tumor differentiation, lymph node metastasis, Lauren classification and invasive depth (all P < 0.05). The positive rate of PAK1 was significantly higher in primary gastric carcinomas than in metastatic lymph nodes (75.0% vs. 52.5%, Chi2 = 4.381, P < 0.05). Survival analysis using the Kaplan-Meier method showed that the expression of PAK1 was a predictor for poor prognosis of the patients with gastric carcinoma (Chi2 = 6.857, P < 0.01). CONCLUSIONS: Expression of PAK1 is an early molecular event in the tumorigenesis of gastric carcinoma. It is also closely correlated the development of gastric carcinoma and the patients' prognosis.

Erythropoietin attenuates vascular calcification by inhibiting endoplasmic reticulum stress in rats with chronic kidney disease.

Previous studies suggested that endoplasmic reticulum (ER) stress induced-apoptosis promoted vascular calcification (VC). Interestingly, erythropoietin (EPO), an endogenous glycoprotein, exerts multiple tissue protective effects by inhibiting ER stress and apoptosis. We investigated the role and potential mechanism of EPO on VC in chronic kidney disease (CKD) rats and cultured vascular smooth muscle cells (VSMCs). The calcification model was established by subtotal nephrectomy in vivo or phosphate overload in vitro. The protein level of EPO receptor (EPOR) was increased in the calcified aortas of CKD rats. EPO prevented the reduction of VSMC phenotypic markers, and reversed the increased calcium content and calcium salt deposition in the aortas of CKD rats and cultured calcified VSMCs. The protein levels of activating transcription factor 4 (ATF4) and glucose-regulated protein 94 (GRP94) were upregulated in aortas and VSMCs under calcifying conditions, indicating ER stress activation. EPO treatment of CKD rats or calcified VSMCs downregulated the protein levels of ATF4 and GRP94. Furthermore, ER stress-mediated apoptosis, determined by the protein levels of CCAAT/enhancer-binding protein-homologous protein and cleaved caspase 12, was increased in tunicamycin or calcification media-treated VSMCs, but the increased effect was reversed in EPO-treated groups. The increased apoptotic cells in calcified VSMCs, as indicated by Hoechst staining and flow cytometry, were downregulated by the co-administration of EPO or 4-phenyl butyric acid. In conclusion, EPO might attenuate VC by inhibiting ER stress mediated apoptosis through EPOR signaling.

Identification of the DNA Replication Regulator MCM Complex Expression and Prognostic Significance in Hepatic Carcinoma.

BACKGROUND: The microliposome maintenance (MCM) complex, MCM2-7, is revealed to be involved in multiple cellular processes and plays a key role in the development and progression of human cancers. However, the MCM complex remains poorly elaborated in hepatic carcinoma (HCC). METHODS: In the study, we found the mRNA and protein level by bioinformatics. We also explored the prognostic value, genetic alteration, interaction network, and functional enrichment of MCM2-7. The MCM expression and correlation among these MCMs in HCC cell lines were identified by western blot. RESULTS: MCM2-7 was significantly increased in HCC tissues compared to normal liver tissues. The high level of MCM2-7 had a positive correlation with poor prognosis. However, MCM2-7 alterations were not correlated with poor OS. MCMs were both increased in HCC cell lines compared to the normal hepatocyte cell line. Furthermore, the positive correlation was found among MCMs in HCC cell lines. CONCLUSIONS: The MCM complex was increased in HCC tissues and cell lines and negatively correlated with prognosis, which might be important biomarkers for HCC.