MicroRNA miR-7-5p targets MARK2 to control metamorphosis in Galeruca daurica.

The MARK2 gene, coding microtubule affinity-regulating kinase or serine/threonine protein kinase, is an important modulator in organism microtubule generation and cell polarity. However, its role in the metamorphosis of insects remains unknown. In this study, we found a conserved miRNA, miR-7-5p, which targets MARK2 to participate in the regulation of the larval-pupal metamorphosis in Galeruca daurica. The dual luciferase reporter assay showed that miR-7-5p interacted with the 3' UTR of MARK2 and repressed its expression. The expression profiling of miR-7-5p and MARK2 displayed an opposite trend during the larval-adult development process. In in-vivo experiments, overexpression of miR-7-5p by injecting miR-7-5p agomir in the final instar larvae down-regulated MARK2 and up-regulated main ecdysone signaling pathway genes including E74, E75, ECR, FTZ-F1 and HR3, which was similar to the results from knockdown of MARK2 by RNAi. In contrast, repression of miR-7-5p by injecting miR-7-5p antagomir obtained opposite effects. Notably, both overexpression and repression of miR-7-5p in the final instar larvae caused abnormal molting and high mortality during the larval-pupal transition, and high mortality during the pupal-adult transition. The 20-hydroxyecdysone (20E) injection experiment showed that 20E up-regulated miR-7-5p whereas down-regulated MARK2. This study reveals that the accurate regulation of miRNAs and their target genes is indispensable for insect metamorphosis.

MicroRNA miR-285 modulates the metamorphosis in Galeruca daurica by targeting Br-C.

BACKGROUND: Galeruca daurica has become a new pest on the Inner Mongolia grasslands since an abrupt outbreak in 2009 caused serious damage. As a pupa indicator during insect metamorphosis, the early response gene of the ecdysone signaling pathway, Broad-Complex (Br-C), plays a vital role in the growth and development of insects. MicroRNAs (miRNAs) are small non-coding RNAs which mediate various biological activities, but it is unknown whether and how Br-C is regulated by miRNAs. RESULTS: Temporal expression profiles revealed that miR-285 and Br-C basically displayed an opposite trend during larval-adult development, and Br-C was sharply up-regulated on the last day of final-instar larvae while miR-285 was significantly down-regulated. Both dual-luciferase reporter assay and miRNA-mRNA interaction assay indicated that miR-285 interacts with the coding sequence of Br-C and represses its expression. Not only overexpression but also downexpression of miR-285 led to the failure of larval to pupal to adult metamorphosis. In addition, both overexpression of miR-285 and silence of Br-C inhibited the expression of Br-C and other ecdysone signaling pathway genes, including E74, E75, ECR, FTZ-F1, and HR3. On the contrary, suppressing miR-285 obtained opposite results. Further experiments showed that 20-hydroxyecdysone down-regulated miR-285 and up-regulated Br-C and above-mentioned genes, whereas juvenile hormone alalogue (JHA) resulted in opposite effects. CONCLUSION: Our results reveal that miR-285 is involved in mediating the metamorphosis in G. daurica by targeting Br-C in the ecdysone signaling pathway. miR-285 and its target Br-C could be as a potential target for G. daurica management. © 2024 Society of Chemical Industry.

Expression Profiling and Functional Analysis of Candidate Odorant Receptors in Galeruca daurica.

Galeruca daurica (Joannis) is an oligophagous pest in the grasslands of Inner Mongolia, China, which feed mainly on Allium spp. Odorant receptors (ORs) play an important role in the olfactory system in insects, and function together with olfactory co-receptor (ORco). In this study, 21 OR genes were identified from the transcriptome database of G. daurica adults, and named GdauOR1-20 and GdauORco. The expression profiles were examined by RT-qPCR and RNA interference (RNAi) and electroantennogram (EAG) experiments were conducted to further identify the olfactory functions of GdauOR4, GdauOR11, GdauOR15, and GdauORco. It was found that 15 GdauORs (OR1, OR3-6, OR8, OR11-13, OR15, OR17-20, and ORco) were mainly expressed in antennae, and the expression levels of GdauORs in adults were affected by age. When GdauOR4, GdauOR15, and GdauORco were silenced by RNAi, the electrophysiological responses to host plant volatiles were significantly decreased in G. daurica. This study lays a necessary foundation for clarifying the mechanism on finding host plants in G. daurica.

Spin switching in tris(8-aminoquinoline)iron(ii)(BPh4)2: quantitative guest-losing dependent spin crossover properties and single-crystal-to-single-crystal transformation.

As a derivative of 2-picolylamine, which contains rich protons favouring hydrogen bond formation to assemble a variety of valuable spin crossover (SCO) compounds, 8-aminoquinoline (aqin) will be a good candidate for constructing new mononuclear bistable state compounds. With the guidance of this view, two solvated compounds [Fe(aqin)3](BPh4)2·2(CH3CN) (1·2CH3CN) and [Fe(aqin)3](BPh4)2·1.5(CH3COCH3) (2·1.5CH3COCH3) were synthesized. The structural characterization and magnetic studies demonstrate that this strategy has been successful. Single-crystal diffraction reveals that both the mononuclear compounds have facial (fac-)-configuration cations, which form hydrogen bonds using -NH2 groups with solvent molecules (acetonitrile or acetone). Subsequent magnetic measurement shows the highly sensitive solvent-dependent occurrence of a spin transition above room temperature for both compounds. Interestingly, for compound 1·2CH3CN, in the successively repeated heating and cooling process, by monitoring the loss of solvent molecules by TGA, the shifting of the spin transition curve is found to be linearly dependent on the fraction of the residual solvent content. Additionally, the desolvated sample can re-solvate with CH3CN and recover the magnetic response reproducibly. Furthermore, after losing the acetonitrile molecules, the single-crystal-to-single-crystal transformation occurred to give 1.

Degradation of Mcl-1 through GSK-3ß Activation Regulates Apoptosis Induced by Bufalin in Non-Small Cell Lung Cancer H1975 Cells.

BACKGROUND/AIMS: Mcl-1, an anti-apoptotic Bcl-2 family member, is often overexpressed in non-small cell lung cancer (NSCLC). Bufalin has been reported to induce apoptosis in various tumor cells. However, there is no report showing that bufalin could downregulate Mcl-1 expression in NSCLC. METHODS: Cell proliferation was analyzed by cell counting kit-8 (CCK-8) assay in H1975 cells. Cell apoptosis was detected by flow cytometry. Mcl-1 mRNA was detected by RT-PCR. The expression of apoptosis-associated proteins in H1975 cells was detected by western blotting. The levels of Mcl-1 ubiquitination and NOXA were analyzed by Immunoprecipitation assay. RESULTS: Cell growth was inhibited by bufalin in a time and dose-dependent manner. Bufalin induced apoptosis in NSCLC cells by activating caspase cascades and downregulating Mcl-1 expression. However, overexpression of Mcl-1 diminished bufalin-induced apoptosis. Furthermore, bufalin did not reduce Mcl-1 mRNA expression in H1975 cells, but strongly promoted Mcl-1 protein degradation. Proteasome inhibitor MG132 markedly prevented the degradation of Mcl-1 and blocked bufalin-induced Mcl-1 reduction. Bufalin did not significantly affect NOXA protein levels, but downregulated the expression of p-GSK-3ß. GSK-3 inhibitor and GSK-3ß siRNA resulted in increased levels of Mcl-1 and reversed the bufalin-induced Mcl-1 degradation. CONCLUSION: Bufalin induced cell apoptosis in H1975 cells may be through downregulation of Mcl-1. Proteasomal degradation of Mcl-1 via GSK-3ß activation was involved in bufalin-induced apoptosis.

Correlations of the expressions of c-Jun and Egr-1 proteins with clinicopathological features and prognosis of patients with nasopharyngeal carcinoma.

This study intended to explore the correlation of the expressions of c-Jun and Egr-1 proteins with clinicopathological features and prognosis of patients with nasopharyngeal carcinoma (NPC). From January 2008 to January 2011, 123 NPC patients and 59 patients with chronic rhinitis were enrolled in this study. Fresh NPC and normal nasopharynx tissue specimens were obtained during surgery. Immunohistochemistry (IHC) was adopted to determine the positive expressions of the c-Jun and Egr-1 proteins. A 5-year clinical follow-up was conducted on all NPC patients. The Kaplan-Meier survival curve and Cox regression model were used for survival analysis. Compared with normal nasopharynx tissues, c-Jun expression was up-regulated but Egr-1 expression was down-regulated in NPC tissues. NPC patients with stage T3-T4 or stage III-IV had higher positive rates of c-Jun expression than those with stage T1-T2 or stage I-II. However, the positive rates of Egr-1 expression was higher in patients with stage T1-T2 or stage III-IV than those with stage T3-T4 or stage I-II. The survival rate of NPC patients with high c-Jun expression was lower than those with low/negative c-Jun expression, while the survival rate of NPC patients with high Egr-1 expression was higher than those with low/negative Egr-1 expression. The Cox regression analysis revealed that stage T3-T4, high c-Jun expression, and low Egr-1 expression were risk factors for poor prognosis of NPC patients. In conclusion, our study suggests that the c-Jun and Egr-1 proteins can serve as novel potential biomarkers for the early diagnosis and prognosis prediction of NPC.

[Analysis on evaluation of medical device adverse events monitoring and some factors influencing MDAE's reporting by medical personnels].

OBJECTIVE: To investigate the knowledge level related to medical device adverse events (MDAEs) among medical personnels, some factors influencing the reporting behavior and supervision and administrative strategies. METHODS: Stratified sampling, cluster sampling and random sampling were adopted together and a questionnaire survey was conducted among 1897 subjects from 33 hospitals. RESULTS: Medical personnels knew a very little about MDAEs and medical devices' post-market monitoring, but their attitudes towards its benefits were positive. Their intentions to report MDAEs were relatively strong, but there were still some barriers about it. CONCLUSION: A monitoring system and a professional training model about MDAEs reporting should be established and improved in hospitals.

[Expression of CD16zeta in NK cells of B-cell non-Hodgkin's lymphoma patients and in vitro killing effect of rituximab combined lymphokine-activated killer cells on B-NHL cells].

BACKGROUND & OBJECTIVE: Natural killer (NK) cells are the main effector of antibody-dependent cellular cytotoxicity (ADCC). The activation disorder of NK cells in cancer patients may affect the treatment effect of monoclonal antibody. Reversing the dysfunction of signal transduction of CD16zeta chain in NK cells and combining lymphokine-activated killer (LAK) cells with rituximab may give rise to synergistic effect. This study was to find out whether the activation disorder of NK cells exist in B-cell non-Hodgkin's lymphoma (B-NHL) patients, whether interleukin-2 (IL-2) can reverse the activation disorder in vitro, and whether the combination of rituximab and LAK cells can produce synergistic antitumor effect. METHODS: Peripheral blood mononuclear cells (PBMCs) were isolated from 69 B-NHL patients and 30 healthy donors by density gradient centrifugation method, and cultured with IL-2 (1 000 U/ml) to prepare LAK cells. The positive rate and median fluorescence intensity (MFI) of CD16zeta chain in PBMCs and LAK cells were detected by flow cytometry (FCM). The expression of CD20 on Raji cells was also detected by FCM. The apoptosis of Raji cells after treatment of rituximab was detected by FCM with Annexin V/PI staining. The cytotoxicity was assessed by lactate dehydrogenase (LDH) release experiment. RESULTS: The positive rate and MFI value of CD16zeta chain on CD56+ cells were significantly lower in B-NHL group than in control group [(63.3+/-16.4)% vs. (97.8+/-3.1)%, P<0.001; 1.3+/-1.3 vs. 3.6+/-1.7, P<0.001]. There was no significant difference in the positive rate and MFI value of CD16zeta on LAK cells between the 2 groups [(99.3+/-4.1)% vs. (99.7+/-3.9)%, P=0.145; 29.2+/-12.5 vs. 31.4+/-13.8, P=0.44]. At the concentration of 40 mug/ml, rituximab completely combined CD20 antigens on cell membrane. The obvious enhancive effect of rituximab on cell apoptosis appeared at 24 h after treatment. The killing rate of Raji cells was significantly higher in rituximab combined LAK group than in LAK group (P<0.05). While the combination of LAK cells and Herceptin (40 mug/ml) didn't make a significant increase as compared with Herceptin alone (P>0.05). There was no significant difference in killing rate of Jurket cells between rituximab combined LAK group and LAK group (P>0.05). CONCLUSIONS: The down-regulation of CD16zeta chain expression widely exists in B-NHL patients, while high dose of IL-2 could enhance the expression of CD16zeta chain greatly in vitro. The combination of rituximab and LAK cells shows strong killing effect on Raji cells.