Expression Of MKK4 Research Articles

Protective effect of Tongqiao Huoxue Decoction containing cerebrospinal fluid on OGD/R-damaged HT22 cells via regulation of ASK1/MKK4/JNK signaling pathway

To investigate the protective effect of Tongqiao Huoxue Decoction containing cerebrospinal fluid(TQHXD-CSF) on HT22 cells damaged by oxygen-glucose deprivation/reoxygenation(OGD/R) and whether the mechanism is related to the regulation of ASK1/MKK4/JNK signaling pathway. HT22 cells were subjected to OGD/R to simulate cerebral ischemia-reperfusion injury(CIRI). Then the cells were randomly divided into five groups: blank cerebrospinal fluid(control group), OGD/R group, TQHXD-CSF group, Z-VAD-FMK group(20 μmol·L~(-1)) and TQHXD-CSF+Z-VAD-FMK group. Except the control group, cells in the other groups were reoxygenated for 12 h after 6 h of oxygen and glucose deprivation for modeling OGD/R, and group administration was performed. Cell viability and cytotoxicity were detected by CCK8 and LDH assay kit, respectively and the morphology of HT22 cells was observed by inverted microscope. Western blot and qRT-PCR were employed to detect the protein and mRNA expression levels of Bax, Bcl-2 and caspase-3, respectively. Then HT22 cells were assigned into the control group, OGD/R group, si-NC group, si-ASK1 group, TQHXD-CSF group and TQHXD-CSF+si-ASK1 group. Cell viability, proliferation and apoptosis were determined by CCK8, electric cell-substrate impedance sensing(ECIS), and Hoechst staining and flow cytometry, respectively. The protein expression of MKK4, p-MKK4, JNK, p-JNK, c-Jun, p-c-Jun, Cyt C, Bax, Bcl-2 and caspase-3 was tested by Western blot. The results showed that compared with OGD/R group, TQHXD-CSF significantly enhanced cell viability, improved cell morphology and reduced the protein and mRNA expression levels of Bax, Bcl-2 and caspase-3. In addition, when ASK1 was silenced, compared with OGD/R group, TQHXD-CSF remarkably improved cell viability, and decreased apoptosis rate and the protein expression levels of p-MKK4, p-JNK, p-c-Jun, Cyt C, Bax/Bcl-2 and caspase-3, but the effect was not as good as that of TQHXD-CSF+si-ASK1 group. In conclusion, TQHXD-CSF can inhibit apoptosis mediated by ASK1/MKK4/JNK signaling pathway in OGD/R-damaged HT22 cells, and has protective effect on ischemia-reperfusion injury.

Chlorosis seedling lethality 1 encoding a MAP3K protein is essential for chloroplast development in rice

BackgroundMitogen-activated protein kinase (MAPK) cascades are conserved signaling modules in eukaryotic organisms and play essential roles in immunity and stress responses. However, the role of MAPKs in chloroplast development remains to be evidently established.ResultsIn this study, a rice chlorosis seedling lethality 1 (csl1) mutant with a Zhonghua11 (ZH11, japonica) background was isolated. Seedlings of the mutant were characterized by chlorotic leaves and death after the trefoil stage, and chloroplasts were observed to contain accumulated starch granules. Molecular cloning revealed that OsCSL1 encoded a MAPK kinase kinase22 (MKKK22) targeted to the endoplasmic reticulum (ER), and functional complementation of OsCSL1 was found to restore the normal phenotype in csl1 plants. The CRISPR/Cas9 technology was used for targeted disruption of OsCSL1, and the OsCSL1-Cas9 lines obtained therein exhibited yellow seedlings which phenocopied the csl1 mutant. CSL1/MKKK22 was observed to establish direct interaction with MKK4, and altered expression of MKK1 and MKK4 was detected in the csl1 mutant. Additionally, disruption of OsCSL1 led to reduced expression of chloroplast-associated genes, including chlorophyll biosynthetic genes, plastid-encoded RNA polymerases, nuclear-encoded RNA polymerase, and nuclear-encoded chloroplast genes.ConclusionsThe findings of this study revealed that OsCSL1 played roles in regulating the expression of multiple chloroplast synthesis-related genes, thereby affecting their functions, and leading to wide-ranging defects, including chlorotic seedlings and severely disrupted chloroplasts containing accumulated starch granules.

MYC2 influences salicylic acid biosynthesis and defense against bacterial pathogens in Arabidopsis thaliana.

Arabidopsis MYC2 is a basic helix-loop-helix transcription factor that works both as a negative and positive regulator of light and multiple hormonal signaling pathways, including jasmonic acid and abscisic acid. Recent studies have suggested the role of MYC2 as a negative regulator of salicylic acid (SA)-mediated defense against bacterial pathogens. By using myc2 mutant and constitutively MYC2-expressing plants, we further show that MYC2 also positively influences SA-mediated defense; whereas, myc2 mutant plants are resistant to virulent pathogens only, MYC2 over-expressing plants are hyper-resistant to multiple virulent and avirulent strains of bacterial pathogens. MYC2 promotes pathogen-induced callose deposition, SA biosynthesis, expression of PR1 gene, and SA-responsiveness. Using bacterially produced MYC2 protein in electrophoretic mobility shift assay (EMSA), we have shown that MYC2 binds to the promoter of several important defense regulators, including PEPR1, MKK4, RIN4, and the second intron of ICS1. MYC2 positively regulates the expression of RIN4, MKK4, and ICS1; however, it negatively regulates the expression of PEPR1. Pathogen inoculation enhances MYC2 association at ICS1 intron and RIN4 promoter. Mutations of MYC2 binding site at ICS1 intron or RIN4 promoter abolish the associated GUS reporter expression. Hyper-resistance of MYC2 over-expressing plants is largely light-dependent, which is in agreement with the role of MYC2 in SA biosynthesis. The results altogether demonstrate that MYC2 possesses dual regulatory roles in SA biosynthesis, SA signaling, pattern-triggered immunity (PTI), and effector-triggered immunity (ETI) in Arabidopsis.

Electroacupuncture Pretreatment Regulates Apoptosis of Myocardial Ischemia-Reperfusion Injury in Rats Through RhoA/p38MAPK Pathway Mediated by miR-133a-5p.

The electroacupuncture (EA) pretreatment possesses a beneficial effect on myocardial ischemia/reperfusion (I/R) injury. However, the molecular mechanism of the EA effect is not fully understood. The study aimed to explore the protective effect of EA pretreatment on myocardial ischemia-reperfusion injury (MIRI) and apoptosis-related mechanisms in rats. Rats underwent in vivo myocardial ischemia-reperfusion, EA pretreatment, or intravenous injection of antagomirs. Cardiac function, infarct area, and myocardial cell apoptosis were measured. Meanwhile, the expressions of MKK3, MKK6, p38MAPK, Bax, Bcl-2, and Caspase-3 were also detected. We found that EA pretreatment significantly reduced infarct area and myocarpal cell apoptosis and enhanced cardiac function. EA pretreatment decreased the expression of Bax, Caspase-3, MKK3, MKK6, p38MAPK, Bax, and Caspase-3. In conclusion, The EA pretreatment down regulated the expression of MKK3, MKK6, and p38MAPK through the RhoA/p38MAPK pathway. EA pretreatment protect MIRI rats from apoptosis by down regulating the expression of MKK3, MKK6, and p38MAPK, thereby reducing the expression of Bax, Caspase-3 and up regulating the expression of Bcl-2, which mechanism is closely related to the RhoA/p38MAPK pathway mediated by miR-133a-5p.

BAFF, involved in B cell activation through the NF-κB pathway, is related to disease activity and bone destruction in rheumatoid arthritis.

B cell activating factor of TNF family (BAFF) is a member of TNF ligand superfamily and plays a key role in B cell homeostasis, proliferation, maturation, and survival. In this study, we detected BAFF level, the expressions of BAFF receptors and important molecules in NF-κB pathway in rheumatoid arthritis (RA) patients and analyzed the correlation between BAFF level and clinical variables, laboratory parameters or X-ray scores in order to elucidate the roles of BAFF in RA. A total of 50 RA patients and 50 healthy controls (HCs) were enrolled. We showed that the serum BAFF level in RA patients was significantly higher than that of HCs, and the percentages of B cell subsets (CD19+ B cells, CD19+CD27+ B cells, CD19+CD20+CD27+ B cells, and CD19+CD20-CD27+ B cells) in the serum of RA patients were significantly increased compared with those of HCs. The percentages of CD19+BAFFR+ B cells, CD19+ BCMA+ B cells, and CD19+ TACI+ B cells in RA patients were significantly increased compared with those in HCs. The expression of important molecules in the NF-κB pathway (MKK3, MKK6, p-P38, p-P65, TRAF2, and p52) was significantly higher in RA patients than in HCs, but p100 level in RA patients was lower than that in HCs. The serum BAFF level was positively correlated with C-reactive protein, rheumatoid factor, disease activity score (in 28 joints), swollen joint counts, tender joint counts, and X-ray scores. When normal B cells were treated with BAFF in vitro, the percentages of the B cell subset and the expression of BAFF receptors were significantly upregulated. BAFF also promoted the expression of MKK3, MKK6, p-P38, p-P65, TRAF2, and p52. In conclusion, this study demonstrates that BAFF level is correlated with the disease activity and bone destruction of RA. BAFF is involved in the differentiation, proliferation, and activation of B cells in RA through NF-κB signaling pathway, suggesting that BAFF might be an ideal therapeutic target for RA.

Nonylphenol Induces Apoptosis through ROS/JNK Signaling in a Spermatogonia Cell Line.

Nonylphenol (NP) is an endocrine-disruptor chemical that negatively affects reproductive health. Testes exposure to NP results in testicular structure disruption and a reduction in testicular size and testosterone levels. However, the effects of NP on spermatogonia in testes have not been fully elucidated. In this study, the molecular mechanisms of NP in GC-1 spermatogonia (spg) cells were investigated. We found that cell viability significantly decreased and apoptosis increased in a dose-dependent manner when GC-1 spg cells were exposed to NP. Furthermore, the expression levels of the pro-apoptotic proteins increased, whereas anti-apoptosis markers decreased in NP-exposed GC-1 spg cells. We also found that NP increased reactive oxygen species (ROS) generation, suggesting that ROS-induced activation of the MAPK signaling pathway is the molecular mechanism of NP-induced apoptosis in GC-1 spg cells. Thus, NP could induce c-Jun phosphorylation; dose-dependent expression of JNK, MKK4, p53, and p38; and the subsequent inhibition of ERK1/2 and MEK1/2 phosphorylation. The genes involved in apoptosis and JNK signaling were also upregulated in GC-1 spg cells treated with NP compared to those in the controls. Our findings suggest that NP induces apoptosis through ROS/JNK signaling in GC-1 spg cells.

A novel triptolide derivative ZT01 exerts anti-inflammatory effects by targeting TAK1 to prevent macrophage polarization into pro-inflammatory phenotype

Sepsis is a main reason for death in intensive care units, inflammation is closely related to sepsis. Anti-inflammation plays an important role in treating of sepsis. ZT01 is a triptolide derivative with strong anti-inflammatory activity and low toxicity. The purpose of this study is to evaluate the anti-inflammatory activity of ZT01 under the sepsis condition and explore the underlying molecular mechanisms. Two in vivo model of sepsis, caecal ligation and puncture or intraperitoneal injection of LPS in C57BL/6, were used to evaluate the therapeutic effects of ZT01. In vitro, the anti-inflammatory properties of ZT01 were assessed in IFN-γ or LPS-induced macrophages by ELISA, RT-PCR, western blotting and co-immunoprecipitation. Macrophages were used to investigate the polarization phenotype by flow cytometry. The results showed, ZT01 significantly attenuated inflammatory response of sepsis in serum or lung tissue by inhibiting production of pro-inflammatory factors and improved the survival rate of septic mice in vivo. In cultured macrophages, ZT01 not only decreased the levels of TNF-α and IL-6 but also prevented the TKA1-TAB1 complex formation, thereby inhibiting the phosphorylation expression of MKK4 and JNK, which were all stimulated by LPS. Moreover, ZT01 inhibited the LPS-induced polarization of macrophages into pro-inflammatory phenotype. Adoptive transfer ZT01 pretreated bone marrow-derived macrophages obviously reduced the pro-inflammatory factors in mice after LPS challenge. Our findings suggested that ZT01 exhibited anti-inflammation activity via preventing the pro-inflammatory phenotype of macrophages by blocking the formation of the TAK1-TAB1 complex and subsequently phosphorylation of MKK4 and JNK.

MicroRNA-152 suppresses cell proliferation and tumor growth of bladder cancer by targeting KLF5 and MKK7

B ackground: In bladder cancer, miR-152 has been reported to be downregulated and accompanied by high DNA methylation. However, the molecular mechanisms underlying miR-152 in bladder cancer tumorigenesis remain unknown. M ethods: The expressions of miR-152 in human bladder cancer cell lines (J82, 5637, and T24) and a human bladder epithelial cell line (SV-HUC-1) were compared. The expression of miR-152 was observed after adding DNA methylation inhibitor 5-Aza-dC or after knocking down the major DNA methyltransferase DNMT1. The bladder cancer cell viability was determined using an MTT assay, and the colony formation rate was determined using a colony formation assay. The effects of miR-152 overexpression on cell proliferation and tumor growth were observed in vitro and in vivo. The interactions among miR-152, KLF5, and MKK7 were determined using a luciferase reporter assay. R esults: MiR-152 was downregulated in bladder cancer cell lines in comparison with normal ones. The expression of miR-152 was significantly promoted after adding 5-Aza-dC or after knocking down DNMT1. The overexpression of miR-152 suppressed cell proliferation and tumor growth both in vitro and in vivo. MiR-152 inhibited KLF5 and MKK7 expression by binding with 3’ untranslated regions of their mRNAs. Meanwhile, the overexpression of KLF5 induced MKK7 expression by increasing the transcriptional activity of MKK7. The simultaneous overexpression of miR-152 and KLF5 reversed the inhibition of cell proliferation caused by miR-152 overexpression alone. C onclusions: Our findings suggest that miR-152 acts as a tumor suppressor in bladder cancer, and the overexpression of miR-152 may be a potential strategy for treating bladder cancer. K eywords: cell proliferation, bladder cancer, DNA methylation, miR-152, transcription factor

Expression of MKK7 and Phospho-c-Jun in Glioma and Their Correlation

Expression of MKK7 and Phospho-c-Jun in Glioma and Their Correlation

Correlation betwen MKK4 expression (exp) and clinicopathological features, KRAS/NRAS mutation in metastatic colorectal cancer (mCRC).

e14500 Background: We aimed to evaluate correlation between MKK4 exp & clinicopathological features in mCRC Methods: MKK4 exp was assessed by immunoreactive score (IRS). Staining intensity [0 (no), 1+ (weak), 2+ (moderate), 3+ (strong)] & % of positively stained cells [0 (0%), 1( < 10%), 2 (11-50%), 3 (51-80%), 4 ( > 80%)]. IRS was calculated by this formula (IRS = SI × PP). Cut-offs were explored with ROC analysis. AUC was 0.629 (95%CI:0.419-0.840) & best cut-off value for MKK4 was 2.5 (sensitivity of 68 %). Groups were WIR ('weak'; IRS:0-2) & SIR ('strong'; IRS ≥3). Results: We enrolled 95 patients. Median follow-up was 31.4 months. MKK4 exp strength outcomes are shown in table. K-ras/N-ras mutation rate was 45.2%. WIR patients had longer OS (p: 0.03). In WIR group, metastatic ones at diagnosis had shorter DFS (p:0,05). There was no PFS difference (p: 0,15). In correlation analysis, there was a a negative correlation between MKK4 expression & 1st line treatment resistance, k-ras/n-ras mt, OS, PFS, DFS (r: -0.01; r: -0,06; -0,02; -0,10; -0,06; -0,34) ( p:0,04 for DFS). Conclusions: MKK4 exp inversely correlates with survival outcomes & ras mutation. WIR leads longer DFS & OS. [Table: see text]

MKK7 transcription positively or negatively regulated by SP1 and KLF5 depends on HDAC4 activity in glioma.

JNK activity has been implicated in the malignant proliferation, invasion and drug-resistance of glioma cells (GCs), but the molecular mechanisms underlying JNK activation are currently unknown. Here, we reported that MKK7, not MKK4, directly activates JNK in GCs and exerts oncogenic effects on tumor formation. Notably, MKK7 expression in glioma tissues was closely correlated with the grade of the glioma and JNK/c-Jun activation. Mechanistically, MKK7 transcription critically depends on the complexes formed by HDAC4 and the transcriptional factors SP1 and Krüppel-like factor-5 (KLF5), wherein HDAC4 directly deacetylates both SP1 and KLF5 and synergistically upregulates MKK7 transcription through two SP1 sites located on its promoter. In contrast, the increases in acetylated-SP1 and acetylated-KLF5 after HDAC4 inhibition switched to transcriptionally suppress MKK7. Selective inhibition of HDAC4 by LMK235, siRNAs or blockage of SP1 and KLF5 by the ectopic dominant-negative SP1 greatly reduced the malignant capacity of GCs. Furthermore, suppression of both MKK7 expression and JNK/c-Jun activities was involved in the tumor-growth inhibitory effects induced by LMK235 in U87-xenograft mice. Interestingly, HDAC4 is highly expressed in glioma tissues, and the rate of HDAC4 nuclear import is closely correlated with glioma grade, as well as with MKK7 expression. Collectively, these findings demonstrated that highly expressed MKK7 contributes to JNK/c-Jun signaling-mediated glioma formation. MKK7 transcription, regulated by SP1 and KLF5, critically depends on HDAC4 activity, and inhibition of HDAC4 presents a potential strategy for suppressing the oncogenic roles of MKK7/JNK/c-Jun signaling in GCs.

HPV type 16 E6 and NFX1–123 augment JNK signaling to mediate keratinocyte differentiation and L1 expression

The HPV life cycle is differentiation-dependent, with cellular differentiation driving initiation of the late, productive stage of the viral life cycle. Here, we identify a role for the protein NFX1–123 in regulating keratinocyte differentiation and events of the late HPV life cycle. NFX1–123 itself increased with differentiation of epithelial cells. Greater NFX1–123 augmented differentiation marker expression and JNK phosphorylation in differentiating 16E6-expressing human foreskin keratinocytes (16E6 HFKs). This was associated with altered expression of MKK4 and MKK7, upstream kinase regulators of JNK phosphorylation. Modulating levels of NFX1–123 in HPV16-positive W12E cells recapitulated the effects on differentiation markers, JNK phosphorylation, and MKK4/7 seen in 16E6 HFKs. Crucially, levels of NFX1–123 also correlated with expression of L1, the capsid protein of HPV. Altogether, these studies define a role for NFX1–123 in mediating epithelial differentiation through the JNK signaling pathway, potentially linking expression of cellular genes and HPV genes during differentiation.

Gossypetin is a novel MKK3 and MKK6 inhibitor that suppresses esophageal cancer growth in vitro and in vivo

Gossypetin as a hexahydroxylated flavonoid found in many flowers and Hibiscus. It exerts various pharmacological activities, including antioxidant, antibacterial and anticancer activities. However, the anticancer capacity of gossypetin has not been fully elucidated. In this study, gossypetin was found to inhibit anchorage-dependent and -independent growth of esophageal cancer cells. To identify the molecular target(s) of gossypetin, various signaling protein kinases were screened and results indicate that gossypetin strongly attenuates the MKK3/6-p38 signaling pathway by directly inhibiting MKK3 and MKK6 protein kinase activity in vitro. Mechanistic investigations showed that arginine-61 in MKK6 is critical for binding with gossypetin. Additionally, the inhibition of cell growth by gossypetin is dependent on the expression of MKK3 and MKK6. Gossypetin caused G2 phase cell cycle arrest and induced intrinsic apoptosis by activating caspases 3 and 7 and increasing the expression of BAX and cytochrome c. Notably, gossypetin suppressed patient-derived esophageal xenograft tumor growth in an in vivo mouse model. Our findings suggest that gossypetin is an MKK3 and MKK6 inhibitor that could be useful for preventing or treating esophageal cancer.

Cordyceps militaris Extract Inhibits the NF-κB pathway and Induces Apoptosis through MKK7-JNK Signaling Activation in TK-10 Human Renal Cell Carcinoma

The ubiquitous transcription factor, NF-κB, has been reported to inhibit apoptosis and induce drug resistance in cancer cells. Cordyceps militaris extract (CME) is involved in the regulation of the NF-κB signaling pathway. However, the detailed role of CME in the suppression of the NF-κB signaling pathway is unclear. We found that CME dose-dependently inhibited tumor necrosis factor-α (TNF-α)-induced NF-κB activation in TK-10 human renal cell carcinoma. CME prevented NF-κB from translocating to the nucleus, which resulted in the downregulation of GADD45B, upregulation of MKK7, and phosphorylation of JNK (p-JNK). The increased activation of Bax led to pronounced CME-induced apoptosis, which occurred through caspase-3. Furthermore, the siRNA-mediated knockdown of GADD45B inhibited MKK7 expression, whereas the siRNA-mediated inhibition of MKK7 downregulated p-JNK and the JNK inhibitor, SP600125, inhibited Bax expression. Thus, these results indicated that CME inhibited the activation of GADD45B via the inhibition of NF-κB activation, which upregulated the MKK7-JNK signaling pathway to induce apoptosis in TK-10 cells. Thus, this study reveals a novel anticancer function of CME.

CP-25, a Novel Anti-inflammatory and Immunomodulatory Drug, Inhibits the Functions of Activated Human B Cells through Regulating BAFF and TNF-alpha Signaling and Comparative Efficacy with Biological Agents.

Paeoniflorin-6′-O-benzene sulfonate (code: CP-25) was the chemistry structural modifications of Paeoniflorin (Pae). CP-25 inhibited B cells proliferation stimulated by B cell activating factor belonging to the TNF family (BAFF) or Tumor necrosis factor alpha (TNF-alpha). CP-25, Rituximab and Etanercept reduced the percentage and numbers of CD19+ B cells, CD19+CD20+ B cells, CD19+CD27+ B cells and CD19+CD20+CD27+ B cells induced by BAFF or TNF-alpha. There was significant difference between CP-25 and Rituximab or CP-25 and Etanercept. CP-25 down-regulated the high expression of BAFFR, BCMA, and TACI stimulated by BAFF or TNF-alpha. The effects of Rituximab and Etanercept on BAFFR or BCMA were stronger than that of CP-25. CP-25, Rituximab and Etanercept down-regulated significantly the expression of TNFR1 and TNFR2 on B cell stimulated by BAFF or TNF-alpha. CP-25, Rituximab and Etanercept down-regulated the expression of MKK3, P-p38, P-p65, TRAF2, and p52 in B cells stimulated by BAFF and the expression of TRAF2 and P-p65 in B cells stimulated by TNF-alpha. These results suggest that CP-25 regulated moderately activated B cells function by regulating the classical and alternative NF-κB signaling pathway mediated by BAFF and TNF-alpha-TRAF2-NF-κB signaling pathway. This study suggests that CP-25 may be a promising anti-inflammatory immune and soft regulation drug.

A plasma mir-125a-5p as a novel biomarker for Kawasaki disease and induces apoptosis in HUVECs.

BackgroundKawasaki disease (KD) is a childhood systemic vasculitis that exhibits a specific preference for the coronary arteries. The aetiology remains unknown and there are no especially diagnostic tests. microRNAs (miRNAs) are 18 to 23 nucleotides non-coding RNAs that are negative regulator of gene expression and play a crucial role in the regulatory network of the genome. Recently, circulating miRNAs have been found presentation in human plasma and displayed some characteristics of the ideal biomarker. However, few researches explored differentially expressed miRNAs in the plasma of KD patients. Our study is to identify circulating miRNAs in KD plasma which can serve as potential biomarkers of KD diagnosis.Materials and methodsThe total of five pairs of acute KD and normal plasma samples were analyzed using ABI miRNAs TLDA Assay chip. Differentially expression of miR-125a-5p in plasma were confirmed by quantitative real-time PCR (qRT-PCR) in independent cohort (acute KD = 30, convalescent KD = 30 and healthy control = 32). After bioinformatics prediction, miR-125a-5p vector and inhibitor were transfected into HUVECs respectively, to observe MKK7 expression as a potential target gene. Flow cytometry was used to analyze apoptosis. The mRNA and protein levels of desired genes including MKK7, Caspase-3, Bax and Bcl2 were detected by qRT-PCR and western blotting.ResultsEighteen miRNAs were differentially expressed in acute KD’s plasma compared with healthy control. miR-125a-5p was significantly increased in plasma of KD patients (p = 0.000), but no variation between acute and convalescent KD (p = 0.357). Moreover, the results from the gain and loss functions of miR-125a-5p in HUVECs have shown that miR-125a-5p remarkably suppressed MKK7 expression, as a novel target gene. Importantly, miR-125a-5p also induced apoptosis in HUVECs through inhibition MKK7 levels to regulate Bax/Bcl2 pathway resulting to activate Caspase-3.ConclusionOur study indicated that the circulating miR-125a-5p levels in KD’s plasma have remarkably evaluated compared with healthy individuals. miR-125a-5p might play a role in the development of KD by regulating target gene MKK7 to induce apoptosis in vascular endothelial cells. Therefore, our findings have suggested that detected miR-125a-5p levels in plasma could be used as a potential biomarker in early KD diagnosis.

L1 Cell Adhesion Molecule Promotes Migration and Invasion via JNK Activation in Extrahepatic Cholangiocarcinoma Cells with Activating KRAS Mutation.

Extrahepatic cholangiocarcinoma (ECC), a malignant tumor of biliary origin, has a poor prognosis with limited treatment options. The KRAS oncogene is the most commonly mutated gene in ECC and one of the factors that predicts a poor prognosis and low survival rate. L1 cell adhesion molecule (L1CAM) is expressed in ECC cells and acts as an independent poor prognostic factor in predicting patient survival. In this study we investigate the functional significance of L1CAM in ECC cells with activating KRAS mutation. We selected an ECC cell line, EGI-1, with activating KRAS mutation, and then confirmed its expression of L1CAM by RT-PCR, western blot analysis, and flow cytometry. The suppression of L1CAM expression (using a specific lentivirus-delivered shRNA) significantly decreased the migratory and invasive properties of EGI-1 cells, without altering their proliferation or survival. Analyses of signaling effectors in L1CAM-depleted and control EGI-1 cells indicated that L1CAM suppression decreased the levels of both phosphorylated MKK4 and total MKK4, together with c-Jun N-terminal kinase (JNK) phosphorylation. Further, exposure to a JNK inhibitor (SP600125) decreased migration and invasion of EGI-1 cells. These results suggest that L1CAM promotes cellular migration and invasion via the induction of MKK4 expression, leading to JNK activation. Our study is the first to demonstrate a functional role for L1CAM in ECC carrying the activating KRAS mutation. Given that KRAS is the most commonly mutated oncogene in ECC, L1CAM may serve as an attractive therapeutic target for ECC cells with activating KRAS mutation.

Mice haploinsufficient for Map2k7, a gene involved in neurodevelopment and risk for schizophrenia, show impaired attention, a vigilance decrement deficit and unstable cognitive processing in an attentional task: impact of minocycline

RationaleMembers of the c-Jun N-terminal kinase (JNK) family of mitogen-activated protein (MAP) kinases, and the upstream kinase MKK7, have all been strongly linked with synaptic plasticity and with the development of the neocortex. However, the impact of disruption of this pathway on cognitive function is unclear.ObjectiveIn the current study, we test the hypothesis that reduced MKK7 expression is sufficient to cause cognitive impairment.MethodsAttentional function in mice haploinsufficient for Map2k7 (Map2k7+/− mice) was investigated using the five-choice serial reaction time task (5-CSRTT).ResultsOnce stable performance had been achieved, Map2k7+/− mice showed a distinctive attentional deficit, in the form of an increased number of missed responses, accompanied by a more pronounced decrement in performance over time and elevated intra-individual reaction time variability. When performance was reassessed after administration of minocycline—a tetracycline antibiotic currently showing promise for the improvement of attentional deficits in patients with schizophrenia—signs of improvement in attentional performance were detected.ConclusionsOverall, Map2k7 haploinsufficiency causes a distinctive pattern of cognitive impairment strongly suggestive of an inability to sustain attention, in accordance with those seen in psychiatric patients carrying out similar tasks. This may be important for understanding the mechanisms of cognitive dysfunction in clinical populations and highlights the possibility of treating some of these deficits with minocycline.

DUSP1 induces paclitaxel resistance through the regulation of p-glycoprotein expression in human ovarian cancer cells

The heterogeneity and genetic instability of ovarian cancer cells often lead to the development of drug resistance, closely related with the increased cancer-related mortality. In this study, we investigated the role of dual-specificity phosphatase 1 (DUSP1) in the development of the resistance in human ovarian cancer cells against paclitaxel. Overexpression of DUSP1 in HeyA8 human ovarian cancer cells (HeyA8-DUSP1) up-regulated the expression of the drug efflux pump, p-glycoprotein. Consequently, HeyA8-DUSP1 cells are highly resistant to paclitaxel, with the resistance comparable to that of a multi-drug resistance cell line (HeyA8-MDR). Moreover, over expression of DUSP1 significantly increased the activation of p38 MAPK, leaving the activation of ERK1/2 and JNK1/2 unaffected. Pharmacological suppression of p38 MAPK activity prevents the up-regulation of p-glycoprotein expression and the consequent resistance against paclitaxel in HeyA8-DUSP1 cells. By contrast, HeyA8-MDR cells expressed a significantly higher level of DUSP1, but treatment with small interference RNA against DUSP1 significantly suppressed the expression of p-glycoprotein and the resistance against paclitaxel in HeyA8-MDR cells. Ectopic expression of MKK3, an upstream activator of p38 MAPK, significantly up-regulated the expression of p-glycoprotein and increased the consequent resistance against paclitaxel in HeyA8 cells. Collectively, these data indicated that DUSP1 may induce the resistance against paclitaxel through the p38 MAPK-mediated overexpression of p-glycoprotein in human ovarian cancer cells.

META-ANALYSIS OF INTRACORONARY THROMBOLYTIC AS AN ADJUNCT TO PRIMARY PERCUTANEOUS CORONARY INTERVENTION IN ST-SEGMENT ELEVATION MYOCARDIAL INFARCTION

BACKGROUND: We have shown that type I collagen matrix treatment at 3h post-myocardial infarction (MI) leads to less inflammation and improved cardiac function, but the underlying mechanisms remain to be better characterized. MicroRNAs are emerging as key players in the regulation of the post-MI environment. We previously identified that matrix-treated hearts had down-regulated expression of miR-92a, a miRNA with inflammatory effects that is normally upregulated after MI. The goal of this study was to elucidate a possible role of miR-92a in the anti-inflammatory/pro-wound healing effect of matrix treatment post-MI. METHODS/RESULTS: C57BL/6J mice underwent LAD ligation to induce MI. Hearts were removed at 6h, 1d, 2d, 3d, and 7d post-MI and RNA was extracted from infarct and peri-infarct tissue. PCR analysis revealed that miR-92a was initially upregulated at 6h, gradually decreased from 1d to 3d, and then increased again at 7d. The first peak in miR-92a expression coincides with initiation of the inflammatory phase, so we assessed the macrophage response in matrix-treated mice. Three hours post-MI, a separate group of mice received intramyocardial injections of PBS or collagen matrix. Tissues were harvested at 2 days, and miR-92a was confirmed by PCR to be down-regulated in the matrixvs. PBS-treated hearts (p<0.0005). Matrix-treated hearts also had a reduced number of M1 inflammatory macrophages at 2 days (p<0.05). Bone marrow-derived macrophages (BMDMs) were used in vitro to further evaluate matrix regulation of miR-92a-mediated pathways in macrophages. Integrins a5 (ITGa5) and aV (ITGaV), involved in cell-matrix interactions, as well as inflammatory regulators MKK4, CD69, S1PR1 and RAP1b were identified as putative miR-92a targets. BMDMs cultured on the collagen matrix had significantly reduced expression of miR-92a compared to those cultured on TCPS (p<0.001), while the miR-92a target genes integrins a5 and aV were significantly up-regulated. Forced over-expression of miR-92a in matrixcultured BMDMs caused a decrease in the expression of MKK4 (p1⁄40.01), CD69 (p<0.001), ITGa5 (p1⁄40.001), ITGaV (p1⁄40.02), and S1PR1 (p1⁄40.01). Finally, the matrix also enhanced the differentiation of M2 wound healing macrophages as indicated by increased expression of IL-10. CONCLUSION: We report that the beneficial effects of matrix treatment post-MI may be mediated, at least in part, through its ability to regulate miR-92a and pro-wound healing mechanisms in macrophages. These results present the matrix as a novel non-pharmacological approach to locally regulate miRNAs in vivo for reducing inflammation and protecting the myocardium post-MI.