本研究旨在探讨低温等离子体对仔鸡睾丸支持细胞增殖的影响及其调控机制。结果发现，用2.4 W放电功率的低温等离子体间隔6 h处理两次，每次处理时间30 s，对支持细胞的活性、生长速度和细胞的周期进程具有最大的促进作用（P<0.05）。低温等离子体处理增加了睾丸支持细胞线粒体的活性、三磷酸腺苷的产生和呼吸链酶的活性（P<0.05），减少了细胞内活性氧的生成（P<0.05），提高了抗氧化酶的活性（P<0.05），增加了miR-7450-5p的表达（P<0.05），使腺苷一磷酸活化蛋白激酶（adenosine monophosphate-activated protein kinase, AMPK）的水平降低（P<0.05），并且降低了miR-100-5p的表达（P<0.05），使哺乳动物雷帕霉素靶蛋白（mammalian target of rapamycin, mTOR）的水平升高（P<0.05）。支持细胞转染miR-7450-5p抑制剂降低了miR-7450-5p的表达（P<0.05），增加了AMPK的水平（P<0.05），转染miR-100-5p模拟物增加了细胞miR-100-5p的表达（P<0.05），降低了mTOR的水平（P<0.05）。转染miR-7450-5p抑制剂和miR-100-5p模拟物均显著降低了睾丸支持细胞的活性和生长（P<0.05），抑制了细胞周期进程（P<0.05），减少了线粒体的活性、三磷酸腺苷的水平和呼吸链酶的活性（P<0.05），然而低温等离子体处理可以显著改善miR-7450-5p抑制剂和miR-100-5p模拟物对支持细胞增殖的抑制作用（P<0.05）。研究结果表明，低温等离子体处理可能通过影响miRNAs水平与活性氧稳态来调控AMPK-mTOR信号通路，进而通过增加线粒体三磷酸腺苷水平和呼吸链酶活性促进睾丸支持细胞的增殖。本研究优化了低温等离子体促进睾丸支持细胞增殖的处理条件，阐述了其调控的可能机制，为临床实际中利用低温等离子体技术促进睾丸支持细胞增殖奠定基础，从而有利于提高公鸡的繁殖性能。

微RNA (microRNAs,miRNA)、小干扰RNA (small interfering RNA,siRNA) 是真核生物体内触发RNA干扰 (RNA interference,RNAi) 的两种非编码小RNA。两种非编码RNA生物合成通路在黑腹果蝇、埃及伊蚊、家蚕以及其他昆虫中具有广泛研究，但在膜翅目昆虫尤其是寄生蜂中少见。蝶蛹金小蜂是蝴蝶蛹期寄生蜂。本研究通过蝶蛹金小蜂基因组对miRNA、siRNA合成通路鉴定并分析。结果表明除siRNA通路中R2D2、Argonaute-2基因在基因组中分别具有2、3个拷贝，两个通路其他关键基因均只有1个拷贝。结构域保守性分析表明蝶蛹金小蜂相关蛋白与其他物种同源蛋白具有相似结构。对膜翅目昆虫Dicer、Argonaute基因进化分析表明siRNA通路相关基因进化速率更快。但与其他膜翅目昆虫相反，蝶蛹金小蜂中Dicer-2基因进化速率比Dicer-1基因更低。表达分析显示miRNA通路基因在蝶蛹金小蜂成虫中表达量更高而siRNA通路基因表达模式各不相同。本研究为了解寄生蜂miRNA、siRNA生物合成通路及其作用机制提供新的认识。

Methylation of the N6 position of adenine, termed N6-methyladenine, protects DNA from restriction endonucleases via the host-specific restriction-modification system. N6-methyladenine was discovered and has been well studied in bacteria. N6-adenine-specific DNA methyltransferase (N6AMT) is the main enzyme catalyzing the methylation of the adenine base and knowledge of this enzyme was mainly derived from work in prokaryotic models. However, large-scale gene discovery at the genome level in many model organisms indicated that the N6AMT gene also exists in eukaryotes, such as humans, mice, fruit flies and plants. Here, we cloned a N6AMT gene from Nilaparvata lugens (Nlu-N6AMT) and amplified its fulllength transcript. Then, we carried out a systematic investigation of N6AMT in 33 publically available insect genomes, indicating that all studied insects had N6AMT. Genomic structure analysis showed that insect N6AMT has short introns compared with the mammalian homologs. Domain and phylogenetic analysis indicated that insect N6AMT had a conserved N6-adenineMlase domain that is specific to catalyze the adenine methylation. Nlu-N6AMT was highly expressed in the adult female. We knocked down Nlu-N6AMT by feeding dsRNA from the second instar nymph to adult female, inducing retard development of adult female. In all, we provide the first genome-wide analysis of N6AMT in insects and presented the experimental evidence that N6AMT might have important functions in reproductive development and ovary maturation.

Estrogen plays an important role in regulating testicular Sertoli cell number. Furthermore, S-phase kinase-associated protein 2 (SKP2) plays a central role in mammalian cell cycle progression. The objective of this study was to determine whether 17β-estradiol can regulate the expression of SKP2, and the Sertoli cell cycle, via estrogen receptor β (ERβ), the cyclic adenosine monophosphate (cAMP)-protein kinase A (PKA) and extracellular signal-regulated kinase (ERK1/2) pathway. When cultured immature boar Sertoli cells were treated with 17β-estradiol, a time-dependent increase in SKP2 mRNA and protein level was observed by real-time PCR and Western blot, and 17β-estradiol activity peaked at 30 min. Treatment with ICI182780 and ERβ antagonist reduced 17β-estradiol-induced expression of SKP2 and proliferating cell nuclear antigen (PCNA), while increasing the protein concentration of p27kip1. However, the effect of ERa antagonist on these parameters was lower than that of ICI182780 and ERβ. Forskolin had a similar effect as 17β-estradiol on the expression of SKP2, PCNA and p27kip1. Rp-cAMP, H-89 and U0126 treatment reduced 17β-estradiol-induced changes, while H-89 also inhibited ERK1/2 activation. Therefore, 17β-estradiol mainly regulates SKP2 mRNA and protein expression via ERβ-cAMP-PKA and ERK1/2 activation. SKP2 and PCNA expression were positively correlated, while increased SKP2 expression likely resulted in p27kip1 degradation.

A cDNA clone encoding a putative EBF-like protein (DCEBF1) was obtained from total RNA isolated from senescing carnation (Dianthus caryophyllus L.) petals using reverse transcription PCR and rapid-amplification of cDNA ends techniques. The cDNA contained an open reading frame of 1 878 bp corresponding to 625 amino acids. Results of Northern blot indicated DCEBF1 expression was enhanced by endogenous and exogenous ethylene, and was inhibited by STS in petals and ovaries. Upon wounding treatment, DCEBF1 showed a quick increase in mRNA accumulation which was positively correlated with the increase in ethylene production. The levels of DCEBF1 mRNA increased in both petals and ovaries by sucrose treatment compared with the control.