Effect of lncRNA FEZF1-AS1 on viability and apoptosis of LPS-induced vascular endothelial cells by regulating miR-363-3p expression

AIM To investigate the mechanism of long noncoding RNA （lncRNA） FEZF1-AS1 regulating microRNA-363-3p （miR-363-3p） on the viability and apoptosis of lipopolysaocharide （LPS）-induced vascular endothelial cells. METHODS Human umbilical vein endothelial cells （HUVECs） were cultured in vitro. pcDNA-NC, pcDNA-FEZF1-AS1, anti-miR-NC, anti-miR-363-3p, miR-NC and miR-363-3p mimics were transfected into the HUVECs and LPS stimulation was applied for 24 h. RT-qPCR was used to detect the expression of FEZF1-AS1 and miR-363-3p. The cell viability was measured by MTT assay. The apoptotic rate was analyzed by flow cytometry. The dual-luciferase reporter experiment was used to verify the targeted regulation of FEZF1-AS1 and miR-363-3p. Western blot was used to determined the expression of cyclin D1, Ki67 and cleaved caspase-3. RESULTS Compared with control group, the expression level of FEZF1-AS1 in LPS group was significantly reduced （P<0.05）, and the expression level of miR-363-3p was significantly increased （P<0.05）. Compared with pcDNA-NC+LPS group, the cell viability in pcDNA-FEZF1-AS1+LPS group was significantly increased （P<0.05）, the apoptotic rate was significantly reduced （P<0.05）, the protein levels of cyclin D1 and Ki67 were significantly increased （P<0.05）, and the protein level of cleaved caspase-3 was significantly reduced （P<0.05）. Compared with anti-miR-NC+LPS group, the cell viability in anti-miR-363-3p+LPS group was significantly increased （P<0.05）, the apoptotic rate was significantly reduced （P<0.05）, the protein levels of cyclin D1 and Ki67 were significantly increased （P<0.05）, and the protein level of cleaved caspase-3 was significantly reduced （P<0.05）. Dual-luciferase reporter experiment confirmed that FEZF1-AS1 targeted miR-363-3p. Compared with miR-NC+pcDNA-FEZF1-AS1+LPS group, the cell viability in miR-363-3p+pcDNA-FEZF1-AS1+LPS group was significantly reduced （P<0.05）, the apoptotic rate was significantly increased （P<0.05）, the protein levels of cyclin D1 and Ki67 were significantly reduced （P<0.05）, and the protein level of cleaved caspase-3 was significantly increased （P<0.05）. CONCLUSION Over-expression of FEZF1-AS1 promotes the viability and inhibits apoptosis of LPS induced vascular endothelial cells by inhibiting the expression of miR-363-3p.     

Shift of allelochemicals from Sorghum halepense in the soil and their effects on the soil's bacterial community

Johnsongrass (Sorghum halepense [L.]Pers.), an exotic invasive weed in China, secretes the phenolic compounds, p‐hydroxybenzoic acid (p‐HBA) and p‐hydroxybenzaldehyde (p‐HBAL), as the dominant allelochemicals in the root exudates. To better understand how these two allelochemicals affect the soil microbial community in the rhizosphere of S. halepense, the fate of these compounds in the invaded soil and the effect of these phytotoxins on the soil bacterial community were evaluated. The concentrations of the allelochemicals in the soil were determined by a high‐performance liquid chromatography‐ultraviolet/photodiode array after 1, 2, 4, 6, 12 and 24 h of treatment. MiSeq sequencing was undertaken to understand how the bacterial populations in the soil were affected by the allelochemicals. The HPLC results indicated that p‐HBA was degraded by the microorganisms that were present in the soil after 1 h and disappeared after 6 h of incubation. The compound, p‐HBAL, initially was converted to p‐HBA and then the p‐HBA broke down, disappearing after 12 h of incubation in non‐sterile soil. Both p‐HBA and p‐HBAL were stable under sterile soil conditions for up to 24 h. The relative abundance of Proteobacteria was significantly inhibited. However, those of Acidobacteria, Chloroflexi, Verrucomicrobia and Cyanobacteria were increased by the p‐HBAL treatment. These findings suggest that allelochemicals from S. halepense might affect the bacterial community composition in the soil.     

Immunohistochemical Expression of Potential Therapeutic Targets in Canine Thyroid Carcinoma

Background

Thyroid carcinoma is a common endocrine tumor in the dog. Local invasive growth frequently precludes surgical excision and, in up to 38% of dogs, the tumor has already metastasized by the time of diagnosis. Therefore, it is important to investigate new treatment modalities that may be useful for the large number of dogs with inoperable tumors or metastatic disease.

Hypothesis/Objectives

To investigate the immunohistochemical expression of potential therapeutic targets in canine thyroid tumors.

Animals

74 dogs with thyroid neoplasia.

Methods

Immunohistochemistry was performed for thyroglobulin, calcitonin, vascular endothelial growth factor (VEGF), p53, cycloxygenase‐2 (cox‐2), and P‐glycoprotein (P‐gp).

Results

Fifty‐four (73%) tumors were classified as follicular cell thyroid carcinomas (FTCs) and 20 (27%) as medullary thyroid carcinomas (MTCs). Eighty percent of FTCs and all MTCs had a high percentage (76–100%) of neoplastic cells immunopositive for VEGF. Thirteen percent of FTCs and 50% of MTCs expressed cox‐2. Seven percent of FTCs and 70% of MTCs expressed P‐gp. No tumor was immunopositive for p53 expression. Expression of VEGF (P = .034), cox‐2 (P = .013), and P‐gp (P < .001) was significantly higher in MTCs compared to FTCs.

Conclusions and Clinical Importance

VEGF is a potential therapeutic target in both FTC and MTC in dogs. Cox‐2 and P‐gp may be useful molecular targets in canine MTC.     

湖南柑橘衰退病调查分析及弱毒系筛选

 The investigation showed that stem-pitting Citrus tristeza virus (CTV)occurred commonly in citrus production areas in several varieties of Hunan Province. Accurate detection of CTV strains was performed by p23/PCR method, PCR and the results indicated that the most samples were infected with several CTV isolates. Three mild strains were isolated and their pathogenicity was identified by biological identification, it indicated that p23/PCR groups had uniformity with the pathogenicity of CTV isolates. Furthermore, three mild isolates were tested in the cross protection by analysis of biological symptoms and composition of p23 gene. Different protecting effects were observed among these strains and W17 mild isolate was effective.     

Protein expression of cyclin D2 and p16 in proliferation and differentiation of cultured cardiac myocytes

AIM:To investigate the protein expression of cyclin D₂ and p16 in proliferation and differentiation of cultured cardiac myocytes.METHODS:One-day-old Sparague-Dawley rats were used. Cardiac myocytes(CM) were collected by a trypsin-dispersal method and cultured. Cell growth line and fluorescence activated cell sorting (FACS) were used to investigate the proliferation of CM. Ultra-thin sections were made to observe the ultrastructure of CM under transmission electron microscope. The expression of cyclin D₂ and p16 in CM were measured using immunocytochemistry and image analysis.RESULTS:①Results of cell growth line and FACS analysis showed that cultured CM could proliferate in the first 3 cultured days, but the ability decreased quickly, concomitant with differentiation. CM was obseved quiescent in cell cycle three days later. The ultrastructure of CM showed the large amount of myofilaments and mitochondrion. ②The protein expression of cyclin D₂ in 3,4,5 day CM group was 0.89 times(P＜0.05),0.80 times (P＜0.05) and 0.56 times (P＜0.01) of that in 1 day group, respectively. The expression of p16 in CM was increased during the culture process, 2,3,4,5 day group were 1.63 times, 1.72 times, 1.99 times and 2.84 times (P＜0.01) of that in 1 day group, respectively.CONCLUSION:Cultured neonatal rat cardiac myocytes could proliferate during the first 3 days after incubation, but the ability of proliferation decreased, from the fourth day, concomitant with differentiation. Cyclin D₂ and p16 play the key roles in CM postnatal development. Downregulation of cyclin D₂ and upregulation of p16 may induce CM differentiation.     

miR535-3p调控水稻纹枯病抗性的功能研究

 纹枯病是水稻生产中最重要的病害之一,其病原物立枯丝核菌(Rhizoctonia solani)是一种典型的土传真菌。前期我们通过立枯丝核菌侵染水稻后转录组测序,发现多个水稻内源miRNA响应R. solaniAG1-IA的侵染,其中Osa-miR535-3p的表达在R. solani侵染后被抑制。为明确Osa-miR535-3p在水稻应答R. solani侵染中的功能,本研究分别构建了Osa-miR535-3p在感病品种Xudao3背景下的敲除和过表达材料。发现与Xudao3相比,Osa-miR535-3p敲除系的纹枯病发病程度显著加重,而其过表达系的发病程度明显减轻。 Osa-miR535-3p敲除系的二级枝梗数目增多、穗长增加、分蘖数减少、千粒重增加,而其过表达系的株高降低、二级枝梗数目减少、穗长变短、分蘖数增多、千粒重下降,表明Osa-miR535-3p也参与调控水稻的农艺性状。通过RNA-seq通路分析,发现Osa-miR535-3p主要是通过影响植物激素信号通路如乙烯(Ethylene,ET)、吲哚乙酸(3-Indoleacetic acid,IAA)等信号相关基因的表达,调控水稻对纹枯病的抗性。本研究结果将为进一步解析水稻抗纹枯病分子机制提供理论参考,同时将为抗纹枯病水稻品种培育提供新思路。     

The potential of nonpathogenic Fusarium oxysporum and other biological control organisms for suppressing fusarium wilt of banana

The aim of this study was to evaluate the ability of nonpathogenic F. oxysporum and Trichoderma isolates from suppressive soils in South Africa to suppress fusarium wilt of banana in the glasshouse. Several biological control agents and commercial biological control products were included in the study. The isolates were first screened in vitro on potato dextrose agar. In glasshouse evaluations, the fungal and bacterial isolates were established on banana roots before they were replanted in pathogen-infested soil, while the commercial biocontrol agents were applied as directed by the supplier. Banana plantlets were evaluated for disease development after 7 weeks. In vitro tests showed none of the nonpathogenic isolates suppressed Fusarium oxysporum f.sp. cubense ( Foc ), while slight suppression was observed with the two Trichoderma isolates. Results of the glasshouse evaluations revealed that two of the nonpathogenic F. oxysporum isolates, CAV 255 and CAV 241, reduced fusarium wilt incidence by 87·4 and 75·0%, respectively. The known biological control agent Fo47 did not suppress Foc significantly. Pseudomonas fluorescens strain WCS 417, known for its ability to suppress other fusarium wilt diseases (WCS 417), reduced disease incidence by 87·4%. These isolates should be further evaluated for potential application in the field, independently and in combination.