Altered expression of target genes of spinal cord in different itch models compared with capsaicin assessed by RT-qPCR validation

Abstract

Spinal cord plays a central role in the development and progression of pathogenesis of obstinate pruritus. In the current study, four groups of adult male C57Bl/6 mice were investigated; one group treated with saline, while the other groups intradermally injected with compound 48/80, histamine, α-Me-5-HT and capsaicin (algogenic substance), respectively. The intradermal microinjection of pruritic and algogenic compound resulted in a dramatic increase in the itch/algogenic behavior. Analysis of the microarray data showed that 15 genes in spinal cord (C5-C8) were differentially expressed between control group and 48/80 group, in which 9 genes were up-regulated and 6 genes were down-regulated. Furthermore, the results of RT-qPCR validation studies in C5-C8 spinal cord revealed that the 9 mRNA (Sgk1, Bag4, Fos, Ehd2, Edn3, Wdfy, Corin, 4921511E18Rik and 4930423020Rik) showed very different patterns for these different drugs, especially when comparing α-Me-5-HT and capsaicin. In three itch models, Fos and Ehd2 were up-regulated whereas Corin, 4921511E18Rik and 4930423020Rik were down-regulated. Furthermore, Corin and 4930423020Rik were down-regulated in itch model group compared to capsaicin group. Thus the application of microarray technique, coupled with RT-qPCR validation, further explain the mechanism behind itching evoked by pruritic compounds. It can contribute to our understanding of pharmacological methods for prevention or treatment of obstinate pruritus.

Identification of lncRNA and mRNA expression profiles in rat spinal cords at various time‑points following cardiac ischemia/reperfusion

“…It is well known that regulations of gene expression are varied over the time course (64)(65)(66). Our previous research associated with Itchy E3 ubiquitin protein ligase demonstrated that gene expression was significantly different in the C5-C8 spinal cord at 0.5 and 2 h following compound 48/80 injection when compared with the control group (36). Similar to the above method, we also screened key genes in myocardial tissues under 30 min cardiac ischemia following 2 h reperfusion compared with the sham group (67).…”

“…Then the rats were quickly decapitated to limit animal suffering and minimize the effects on the experimental results, and the T1-T4 spinal cord segments were immediately removed and frozen with liquid nitrogen for 1 min, then stored at -80˚C until required. Total RNA from each animal was quantified using a mirVana miRNA Isolation kit (Ambion; Thermo Fisher Scientific, Inc., Waltham, MA, USA), and RNA integrity was assessed according to the manufacturer's protocol, which included standard denaturing agarose gel electrophoresis (35)(36)(37). For RNA-seq, the microarray work was performed by CapitalBio Technology Co. Ltd. (Beijing, China), whereby 6 tissue samples (3 model group samples and 3 control group samples) were used for mRNA and lncRNA microarray analysis (38).…”

“…RT-qPCR analysis. The present study extracted total RNA from the upper thoracic spinal cord segments (T1-T4) (40) using TRIzol ® reagent (Invitrogen; Thermo Fisher Scientific, Inc.) according to our previous research (35)(36)(37). The primers for RT-qPCR were designed based on the lncRNA sequences (Table I), and were synthesized and purified at Invitrogen (Thermo Fisher Scientific, Inc.).…”

Quantitative proteomics reveal the alterations in the spinal�cord after myocardial ischemia‑reperfusion injury in rats

“…Rapid advancements in high-throughput technologies and computational frameworks offer an excellent opportunity to quantify spinal nociception using neuronal activation induced by noxious stimuli. The author's previous study showed that transcriptomics and metabolomics enable the examination of spinal biological systems in unprecedented detail (32)(33)(34)(35)(36). More recently, different patterns were revealed in the metabolic and transcriptional levels of the thoracic spinal cord under myocardial ischemia-reperfusion injury (37)(38)(39).…”

Altered myocardial gene expression profiling in the ischemic tissues at different time points after cardiac ischemia/reperfusion in rats

“…Recently, long non-coding RNAs (lncRNAs) have become an increased research focus that plays critical roles in many biological processes [15][16][17][18][19][20]. Recent studies suggest that lncRNAs are often accompanied by important regulatory functions in the heart [21][22][23].…”