Investigations on the Changes of Serum Proteins in Rabbits after Trimeresurus stejnegeri Venom Injection via Mass Spectrometry-Based Proteomics.

Purpose: There are few studies on protein phosphorylation in the process of snake poisoning. The purpose of this study was to investigate the toxic mechanism of Trimeresurus stejnegeri at the protein level by determining the differential expression of phosphorylated proteins in rabbits after poisoning using proteomics. Methods: The Trimeresurus stejnegeri venom model in rabbits was established by intramuscular injection of 20 mg/kg venom. The serum was collected and the differential expression of phosphorylated proteins in the serum was determined by the iTRAQ technology, TiO2 enriched phosphorylated peptides, and the mass spectrometry analysis. The functional analysis was conducted using ClueGO software and the related mechanism was evaluated by the network analysis of biological interaction. The expression level of related proteins was determined by the Western blotting assay. Results: Compared to the control group, 77 differentially expressed proteins were observed in the model group. These proteins were closely associated with the complement and agglomerate cascade signaling pathways, the HIF signaling pathway, the pentose phosphate pathway, and the cholesterol metabolism signaling pathway. According to the results of network analysis, TF and SCL16A1 were determined as the core proteins, which were identified by the Western blotting assay. Conclusion: The present study provided valuable phosphorylation signal transduction resources for investigating the toxic mechanism and the therapies for Trimeresurus stejnegeri poisoning.

Preliminary study on the mechanism of acupoint injection of bone marrow mesenchymal stem cells in improving blood flow in the rat of hind limb ischemia.

OBJECTIVE: To explore the mechanism of acupoint injection of bone marrow mesenchymal stem cells (BM-MSCs) in improving blood flow in the rat with hind limb ischemia. METHODS: Twenty-four SD rats were randomly divided into 4 groups: normal control group (n = 6), model group (n = 6), BM-MSCs acupoint injection group (AI group, n = 6) and BM-MSC intramuscular injection group (MI group, n = 6). Sanyinjiao (SP 6), Housanli (ST 36), Zhaohai (KI 6), Huantiao (GB 30) and Yanglingquan (GB 34) were selected for the AI group, and five non-acupoints were selected on gastrocnemius and adductor of ischemic hind limbs in the MI group. BM-MSCs were injected to the latter two groups. The rat hind limb ischemia model was established with the method of blocking the femoral artery and its branches. Three weeks after injection of BM-MSCs, in each group, hindlimb adductor and gastrocnemius were taken from the ischemic side. Expressions of vascular endothelial growth factor (VEGF) and transfer growth factor-beta1 (TGF-beta1) in the skeletal muscle were determined with immunohistochemical method, and the small arteries in the skeletal muscle were labeled with alpha-SMA immunohistochemical staining method, the density of small arteries (number of arterioles/number of muscle fibers) and the number of the blood vessel with VEGF positive expression were calculated. The serum levels of VEGF and nitric oxide (NO) were detected. RESULTS: Compared with the model group, the expression of VEGF and TGF-beta1, and the density of small arteries and the number of VEGF-positive blood vessels in the AI group and the MI group significantly increased (both P < 0.01). Compared with the MI group, the density of small arteries and the number of VEGF-positive blood vessels in the AI group significantly increased (both P < 0.01); Compared with the model group and the normal control group, the serum expression quantity of NO and VEGF in the AI group and the MI group were significantly increased (P < 0.01). CONCLUSIONS: Acuppoint injection of BM-MSCs secrets more VEGF, TGF-beta1 and NO to increase angiogenesis and arteriogenesis, so as to improve blood flow of the rats of hind limb ischemic.

[Effect of acupoint injection with bone marrow mesenchymal stem cells on the blood flow in rats with hind limb ischemia].

OBJECTIVE: To investigate the best injection method of the bone marrow mesenchymal stem cells (BM-MSCs) transplantation for the treatment of a rat model with hind limb ischemia. METHODS: Twenty four SD rats with hind limb ischemia were randomly divided into four groups: control group, model group, acupoint BM-MSCs injection group (API group) and thigh muscle BM-MSCs injection group (TMI group). The acupoints of "Sanyinjiao" (SP 6), "Housanli" (ST 36), "Zhaohai" (KI 6), "Huantiao" (GB 30) and "Yanglingquan" (GB 34) were selected for API group, and five non-acupoints were selected on gastrocnemius and adductor of ischemic hind limb for TMI group. Both groups were accepted BM-MSCs transplantion. Model rat with hind limb ischemia was established with the method of blocking the femoral artery and its branches. The changes of blood flow (perfuse unit, PU) was monitored with laser Doppler flowmetry (LDF). In order to describe the visual changes in blood flow, the PU index (PUI) was determined as the ratio of ischemic to non-ischemic hind limb blood perfusion. And also, the levels of VEGF,bFGF in serum were tested to analyze the immunohistochemical expression quantity of VEGF and bFGF. RESULTS: Comparing with the model and the TMI groups, the PUI value on 3rd, 14th and 21th days after BM-MSCs transplantation were significantly increased in the API group (P < 0.05, P < 0.01). In contrast to the model group, the VEGF,bFGF levels in serum and the immunohistochemical expression quantity of VEGF and bFGF in the API and TMI groups were significantly increased (all P < 0.01). CONCLUSION: Transplantation of BM-MSCs through the acupoint can more significantly and quickly increase the blood flow and cause the greater improvement on hind limb ischemia than that of through the way of muscle injection.