MiR-145-5p alleviates hypoxia/reoxygenation- induced cardiac microvascular endothelial cell injury in coronary heart disease by inhibiting Smad4 expression.

OBJECTIVE: To investigate the effects and mechanism of miR-145-5p on hypoxia/reoxygenation (H/R)-induced cardiac microvascular endothelial cell (CMEC) injury in coronary heart disease (CHD). PATIENTS AND METHODS: Patients with CHD (n=96) and healthy volunteers (n=96) were enrolled, and H/R injury model of CMECs was established. The expression of miR-145-5p and mothers against decapentaplegic homolog 4 (Smad4) mRNA in cells was quantified with reverse transcription polymerase chain reaction (RT-PCR). Then, miR-145-5p mimics and Smad4 inhibitor were transfected into CMECs. Cell counting kit-8 (CCK-8) was employed for proliferation detection, flow cytometry for apoptosis detection, and Western Blot for measuring the expression of apoptosis-related proteins and Smad4 protein. RESULTS: The expression of serum miR-145-5p in patients with CHD was significantly lower than that in healthy individuals. The area under the curve (AUC) of miR-145-5p in diagnosing CHD was 0.894, and the expression of miR-145-5p was negatively correlated with that of Smad4 (p<0.05). Over-expression of miR-145-5p promoted the proliferation, inhibited the apoptosis, and reduced inflammatory responses and oxidative stress in H/R-injured CMECs. Moreover, miR-145-5p might negatively regulate the expression of Smad4 in CMECs. Dual-Luciferase reporter assay determined the targeting relation between miR-145-5p and Smad4. CONCLUSIONS: MiR-145-5p is lowly expressed in patients with CHD, and its over-expression effectively alleviates H/R-induced CMEC injury by inhibiting Smad4.

Development of a Simple Hydroponic Assay to Study Vertical and Horizontal Resistance of Soybean and Pathotypes of Phytophthora sojae.

Phytophthora root rot, caused by Phytophthora sojae, is one of the most damaging diseases of soybean and the introgression of Rps (Resistance to P. sojae) genes into elite soybean lines is arguably the best way to manage this disease. Current bioassays to phenotype the gene-for-gene relationship are hampered with respect to reproducibility and long-term stability of isolates, and do not accurately predict horizontal resistance individually. The aim of our study was to investigate a new way of phenotyping P. sojae isolates and vertical and horizontal resistance in soybean that relies on zoospores inoculated directly into a hydroponic system. Inoculation of P. sojae isolates against a set of eight differentials accurately and reproducibly identified pathotypes over a period of two years. When applied to test vertical resistance of soybean lines with known and unknown Rps genes, the bioassay relied on plant dry weight to correctly identify all genes. In addition, simultaneous inoculations of three P. sojae isolates, collectively carrying eight major virulence factors against 64 soybean lines with known and unknown levels of horizontal resistance, separated the plants into five distinct groups of root rot, allowing the discrimination of lines with various degrees of partial resistance. Based on those results, this bioassay offers several advantages in facilitating efforts in breeding soybean for P. sojae resistance and in identifying virulence factors in P. sojae.

Evaluation of Seed and Soil Treatments with Novel Bacillus subtilis Strains for Control of Soybean Root Rot Caused by Fusarium oxysporum and F. graminearum.

Fusarium root rot is an important disease of soybean in Ontario, Canada. This study is to select antagonistic bacterial agents as effective alternatives to chemical pesticides for the control of root rots caused by Fusarium oxysporum and F. graminearum. Twenty-two Bacillus subtilis strains from soybean and corn roots were tested in dual cultures for inhibition of mycelial growth of F. oxysporum and F. graminearum. All strains significantly reduced mycelial growth of F. oxysporum by approximately 17 to 48% and of F. graminearum by 10 to 32%. Ten B. subtilis strains selected based on their larger fungal inhibition zones were evaluated against macroconidial germination. These strains inhibited the spore germination of F. oxysporum by 20 to 48% and of F. graminearum by 14 to 32% in cell-free filtrates. Under greenhouse conditions, the efficacy of seed and soil treatments with B. subtilis strains against the two Fusarium root rot pathogens was evaluated based on root rot severity, seedling emergence, plant height, and root dry weight. Six B. subtilis strains (SB01, SB04, SB23, SB24, SB28, and SB33) from soybean roots and two strains (CB01 and CH22) from corn roots significantly reduced the severity of the two Fusarium root rots in seed or soil treatments. Strains SB01, SB04, SB23, and SB24 were the most effective treatments against both pathogens in either seed or soil treatment. When applied as seed treatments, these four strains reduced root rot severity by 43 to 63% and increased emergence by 13 to 17%, plant height by 9 to 18%, and root dry weight by 8.4 to 19%. When used as soil treatments, they reduced root rot severity by 68 to 74% and increased emergence by 14 to 18%, plant height by 11 to 23%, and root dry weight by 16 to 24%. These results suggest that the novel strains of B. subtilis identified in this research can be effective alternatives to fungicides in managing Fusarium root rots of soybean, and a greater level of efficacy may be achieved when they were used as soil treatments than seed treatments.

Virulence and Genetic Variability Among Isolates of Mycosphaerella pinodes.

Fifty-eight isolates of Mycosphaerella pinodes, collected from western Canada, New Zealand, France, Australia, the United Kingdom, and Ireland, were analyzed for pathogenic and genetic variation according to their virulence on six differential cultivars of field pea (AC Tamor, Bohatyre, Danto, Majoret, Miko, and Radley) and amplified fragment length polymorphism markers. The 56 isolates were classified into 15 pathotypes. Pathotype 1 consisted of 31 isolates that were virulent on all six pea differential cultivars. Pathotypes 14 and 15 consisted of eight isolates that were avirulent on all six differential cultivars or virulent on one of six differential cultivars. The analysis of molecular variance showed that 57.2% of the total variation was caused by differences among populations, and 42.8% was due to molecular diversity within populations. Phylogenetic analysis of molecular variation of isolates showed that most of the Canadian isolates and four Australian isolates formed two clustered groups, respectively, regardless of virulence on the six differential cultivars. Isolates from New Zealand were geographically clustered into two groups. However, the isolates from France, Ireland, and the United Kingdom were clustered with the Canadian isolates.