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Identification and validation of Rab11a in Rat orofacial inflammatory pain model induced by CFA
Miaomiao Liu1, Xin Li2, Jian Wang3
1Department of Respiratory and Critical Care Medicine, Tangdu Hospital of the Fourth Military Medical University, Xi'an, Shaanxi, China.
Abstract:
Orofacial pain (OFP) is a clinically very common and the most troubling condition; however, there is few effective way to relieve OFP. Rab11a, a small molecule guanosine triphosphate enzyme, is one of the Rab member family playing a vital role in intracellular endocytosis and the pain process. Therefore, we investigated the hub genes of rat OFP model induced by Complete Freund's Adjuvant (CFA) via re-analyzing microarray data (GSE111160). We found that Rab11a acted as a key hub gene in the process of OFP. During the validation of Rab11a, the OFP model was established by peripheral injection of CFA, which decreased the head withdrawal threshold (HWT) and head withdrawal lantency (HWL). Rab11a was observed in NeuN of Sp5C instead of GFAP/IBA-1, and double-IF of Rab11a and Fos positive cells were increased on the 7th day after CFA modeling statistically. Rab11a protein expression in TG and Sp5C of CFA group was also significantly increased. Interestingly, injection of Rab11a-targeted short hairpin RNA (Rab11a-shRNA) into Sp5C could reverse the decrease in HWT and HWL and reduce the expression level of Rab11a. Electrophysiological recording further demonstrated that the activity of Sp5C neuron was improved in CFA group, while Rab11a-shRNA considerably decreased the enhancement of Sp5C neuronal activity. Finally, we detected the expression level of p-PI3K, p-AKT, and p-mTOR in Sp5C of rats after injecting the Rab11a-shRNA virus. To our surprise, CFA upregulated the phosphorylation of PI3K, AKT and mTOR in Sp5C, and Rab11a-shRNA downregulated these molecules' expression. Our data suggest that CFA activates the PI3K/AKT signaling pathway through up-regulating Rab11a expression, which can induce OFP hyperalgesia development furtherly. Targeting Rab11a may be a novel treatment strategy for OFP.
Insights
Rab11a is identified as a key gene in orofacial pain (OFP). Targeting Rab11a with Rab11a-shRNA reversed pain behaviors and modulated the PI3K/AKT pathway, suggesting Rab11a as a potential OFP treatment target.
Area of Science:
- Neuroscience
- Molecular Biology
- Pain Research
Background:
- Orofacial pain (OFP) is a common and debilitating condition with limited effective treatments.
- Rab11a, a Rab GTPase, is implicated in intracellular trafficking and pain pathways.
- Identifying key molecular players in OFP is crucial for developing novel therapeutic strategies.
Purpose of the Study:
- To investigate the role of Rab11a as a hub gene in a rat model of orofacial pain (OFP).
- To validate Rab11a's involvement in OFP development and its potential as a therapeutic target.
Main Methods:
- Re-analysis of microarray data (GSE111160) to identify hub genes in a Complete Freund's Adjuvant (CFA)-induced OFP rat model.
- Validation using behavioral tests (head withdrawal threshold and latency), immunohistochemistry, Western blotting, and electrophysiological recordings.
- Intervention with Rab11a-targeted short hairpin RNA (Rab11a-shRNA) delivered to the Sp5C region.
Main Results:
- Rab11a was identified as a key hub gene in the OFP model.
- CFA injection induced OFP behaviors and increased Rab11a expression in the trigeminal nucleus (Sp5C).
- Rab11a-shRNA treatment reversed pain behaviors, reduced Rab11a expression, normalized Sp5C neuronal activity, and downregulated the PI3K/AKT/mTOR pathway.
Conclusions:
- Rab11a plays a critical role in the development of CFA-induced orofacial pain.
- The PI3K/AKT signaling pathway is activated by Rab11a in OFP.
- Targeting Rab11a represents a promising novel therapeutic strategy for managing orofacial pain.
