Rational design of microRNA–siRNA chimeras for multifunctional target suppression

MicroRNAs (miRNAs) are involved in a variety of human diseases by simultaneously suppressing many gene targets. Thus, the therapeutic value of miRNAs has been intensely studied. However, there are potential limitations with miRNA-based therapeutics such as a relatively moderate impact on gene target regulation and cellular phenotypic control. To address these issues, we proposed to design new chimeric small RNAs (aiRNAs) by incorporating sequences from both miRNAs and siRNAs. These aiRNAs not only inherited functions from natural miRNAs, but also gained new functions of gene knockdown in an siRNA-like fashion. The improved efficacy of multifunctional aiRNAs was demonstrated in our study by design and testing of an aiRNA that inherited the functions of both miR-200a and an AKT1-targeting siRNA for simultaneous suppression of cancer cell motility and proliferation. The general principles of aiRNA design were further validated by engineering new aiRNAs mimicking another miRNA, miR-9. By regulating multiple cellular functions, aiRNAs could be used as an improved tool over miRNAs to target disease-related genes, thus alleviating our dependency on a limited number of miRNAs for the development of RNAi-based therapeutics.     

Cytosolic phospholipase A2α sustains pAKT,pERK and AR levels in PTEN-null/mutated prostate cancer cells

Constitutive phosphorylation of protein kinase B (AKT) is a common feature of cancer caused by genetic alteration in the phosphatase and tensin homolog (PTEN) gene and is associated with poor prognosis. This study determined the role of cytosolic phospholipase A₂α (cPLA₂α) in AKT, extracellular signal-regulated kinase (ERK) and androgen receptor (AR) signaling in PTEN-null/mutated prostate cancer cells. Doxycycline (Dox)-induced expression of cPLA₂α led to an increase in pAKT, pGSK3β and cyclin D1 levels in LNCaP cells that possess a PTEN frame-shift mutation. In contrast, silencing cPLA₂α expression with siRNA decreased pAKT, pGSK3β and cyclin D1 levels in both PC-3 (PTEN deletion) and LNCaP cells. Silencing of cPLA₂α decreased pERK and AR protein levels. The inhibitory effect of cPLA₂α siRNA on pAKT and AR protein levels was reduced by the addition of arachidonic acid (AA), whereas the stimulatory effect of AA on pAKT, pERK and AR levels was decreased by an inhibitor of 5-hydroxyeicosatetraenoic acid production. Pharmacological blockade of cPLA₂α with Efipladib reduced pAKT and AR levels with a concomitant inhibition of PC-3 and LNCaP cell proliferation. These results demonstrate an important role for cPLA₂α in sustaining AKT, ERK and AR signaling in PTEN-null/mutated prostate cancer cells and provide a potential molecular target for treating prostate cancer.     

Defence responses regulated by jasmonate and delayed senescence caused by ethylene receptor mutation contribute to the tolerance of petunia to Botrytis cinerea

Ethylene and jasmonate (JA) have powerful effects when plants are challenged by pathogens. The inducible promoter‐regulated expression of the Arabidopsis ethylene receptor mutant ethylene‐insensitive1‐1 (etr1‐1) causes ethylene insensitivity in petunia. To investigate the molecular mechanisms involved in transgenic petunia responses to Botrytis cinerea related to the ethylene and JA pathways, etr1‐1‐expressing petunia plants were inoculated with Botrytis cinerea. The induced expression of etr1‐1 by a chemical inducer dexamethasone resulted in retarded senescence and reduced disease symptoms on detached leaves and flowers or intact plants. The extent of decreased disease symptoms correlated positively with etr1‐1 expression. The JA pathway, independent of the ethylene pathway, activated petunia ethylene response factor (PhERF) expression and consequent defence‐related gene expression. These results demonstrate that ethylene induced by biotic stress influences senescence, and that JA in combination with delayed senescence by etr1‐1 expression alters tolerance to pathogens.     

MACC1 Down-Regulation Inhibits Proliferation and Tumourigenicity of Nasopharyngeal Carcinoma Cells through Akt/β-Catenin Signaling Pathway

The present study was aimed at investigating the expression of metastasis-associated in colon cancer 1 (MACC1) in nasopharyngeal carcinoma (NPC), its relationship with β-catenin, Met expression and the clinicopathological features of NPC, and its roles in carcinogenesis of NPC. Our results showed that MACC1 expression was higher in NPC cells and tissues than that in normal nasopharyngeal cells and chronic inflammation of the nasopharynx tissues, respectively. MACC1 expression was closely related to the clinical stage (p = 0.005) and the N classification (p<0.05) of NPC. Significant correlations between MACC1 expression and Met expression (p = 0.003), MACC1 expression and β-catenin abnormal expression (p = 0.033) were found in NPC tissues. MACC1 knockdown dramatically inhibited cellular proliferation, migration, invasion, and colony formation, but induced apoptosis in NPC cells compared with the control group. Furthermore, MACC1 down-regulation inhibited phosphorylated-Akt (Ser473) and β-catenin expression in NPC cells, but phosphorylated-Erk1/2 expression was not altered. Further study showed that phosphotidylinsitol-3-kinase inhibitor downregulated β-catenin and Met expression in NPC cells. There was a significant relationship between MACC1 expression and phosphorylated-Akt expression (p = 0.03), β-catenin abnormal expression and phosphorylated-Akt expression (p = 0.012) in NPC tissue, respectively. In addition, Epstein Barr virus-encoded oncogene latent membrane protein 1 upregulated MACC1 expression in NPC cells. Our results firstly suggest that MACC1 plays an important role in carcinogenesis of NPC through Akt/β-catenin signaling pathway. Targeting MACC1 may be a novel therapeutic strategy for NPC.     

The Shape Trail Test: Application of a New Variant of the Trail Making Test

Objective

The Trail making test (TMT) is culture-loaded because of reliance on the Latin alphabet, limiting its application in Eastern populations. The Shape Trail Test (STT) has been developed as a new variant. This study is to examine the applicability of the STT in a senile Chinese population and to evaluate its potential advantages and disadvantages.

Method

A total of 2470 participants were recruited, including 1151 cognitively normal control (NC), 898 amnestic mild cognitive impairment (aMCI), and 421 mild Alzheimer disease (AD) patients. Besides the STT, the Mini mental state examination and a comprehensive neuropsychological battery involving memory, language, attention, executive function and visuospatial ability were administered to all the participants. In a subgroup of 100 NC and 50 AD patients, both the STT and the Color Trail Test (CTT) were performed.

Results

In NC, the time consumed for Part A and B (STT-A and STT-B) significantly correlated with age and negatively correlated with education (p<0.01). STT-A and B significantly differed among the AD, aMCI and NC. The number that successfully connected within one minute in Part B (STT-B-1 min) correlated well with STT-B (r = 0.71, p<0.01) and distinguished well among NC, aMCI and AD. In the receiver operating characteristic curve analysis, the AUCs (area under the curve) for STT-A, STT-B, and STT-B-1min in identifying AD were 0.698, 0.694 and 0.709, respectively. The STT correlated with the CTT, but the time for completion was longer.

Conclusion

The TMT is a sensitive test of visual search and sequencing. The STT is a meaningful attempt to develop a “culture-fair” variant of the TMT in addition to the CTT.     

Changes in heparan sulfate sulfotransferases and cell-surface heparan sulfate during SKM-1 cells granulocytic differentiation and A549 cells epithelial-mesenchymal transition

Glycoconjugate Journal - Heparan sulfate (HS) with various sulfation patterns is one of important modulators of cancer cell fate through interacting with numerous growth factors. Here we found HS...     

miR-222 inhibits cardiac fibrosis in diabetic mice heart via regulating Wnt/β-catenin-mediated endothelium to mesenchymal transition

miR-222 participates in many cardiovascular diseases, but its effect on cardiac remodeling induced by diabetes is unclear. This study evaluated the functional role of miR-222 in cardiac fibrosis in diabetic mice. Streptozotocin (STZ) was used to establish a type 1 diabetic mouse model. After 10 weeks of STZ injection, mice were intravenously injected with Ad-miR-222 to induce the overexpression of miR-222. miR-222 overexpression reduced cardiac fibrosis and improved cardiac function in diabetic mice. Mechanistically, miR-222 inhibited the endothelium to mesenchymal transition (EndMT) in diabetic mouse hearts. Mouse heart fibroblasts and endothelial cells were isolated and cultured with high glucose (HG). An miR-222 mimic did not affect HG-induced fibroblast activation and function but did suppress the HG-induced EndMT process. The antagonism of miR-222 by antagomir inhibited HG-induced EndMT. miR-222 regulated the promoter region of β-catenin, thus negatively regulating the Wnt/β-catenin pathway, which was confirmed by β-catenin siRNA. Taken together, our results indicated that miR-222 inhibited cardiac fibrosis in diabetic mice via negatively regulating Wnt/β-catenin-mediated EndMT.     

The correlation between CT features and insulin resistance levels in patients with T2DM complicated with primary pulmonary tuberculosis

The aim is to investigate the correlation between computed tomography (CT) features and insulin resistance levels in patients with type 2 diabetes mellitus (T2DM) complicated with primary pulmonary tuberculosis (PTB). Nearly, 268 untreated PTB patients complicated with T2DM were divided into two groups according to the optimal cutoff value of HOMA-IR score for the Chinese population: HOMA-IR ≤ 2.69 (Group I: 74 patients), >2.69 (Group II: 194 patients). The basic characteristics and changes of CT manifestations were analyzed. In the two groups, the detection rate of large segmented leafy shadow was 39.2% and 78.9%; the air bronchogram sign detection rate was 40.5% and 80.9%; the discovery rate of mouth-eaten cavity was 33.8% and 73.7%; the thin-walled cavity detection rate was 2.7% and 16.0%; the rate of multiple cavities was 35.1% and 69.6%; and bronchial tuberculosis was found in 4.1% and 35.6%, respectively. The detection rates of lesions in Group II were significantly higher than in Group I (p < .05). HOMA-IR was found independently associated with large segmented leafy shadow, air bronchial sign, thin-walled cavity, and bronchial tuberculosis. The level of insulin resistance can effectively reflect the severity of PTB patients with T2DM. CT scan can directly provide image information in clinics. These two examinations can guide clinicians to accurately formulate subsequent treatment plans.