ObjectiveTo construct a cationic microbubble (CMB), and investigate the enhancement of gene transfection efficiency and therapeutic effect of ultrasound-targeted microbubble destruction (UTMD) in vivo with CMB compared to definity MB (DMB).Methods In vitro, the CMB was prepared by the method of thin film hydration. The morphology, size, zeta potential, and gene-carrying capacity of CMB were compared with the DMB. In vivo, the firefly luciferase gene which was used as a reporter gene was targeted transfected into myocardium of 16 rats with CMB and DMB, respectively. The gene transfection efficiency and targeting were observed dynamically. Then, ischemia-reperfusion (I/R) model was performed on 64 rats. The models of 60 rats were successfully confirmed by using ultrasonography at 5 days after I/R. The rats were divided into 3 groups (n=20) randomly. The control group received DMB carrying empty plasmid for transfection; DMB group received DMB carrying AKT plasmid for transfection; and CMB group received CMB carrying AKT plasmid for transfection. The cardiac perfusion, cardiac function, infarct size, and infarct thickness were measured by ultrasonography and histological observations after treatment. In addition, the capillary and arteriolar densities were measured with immunohistochemical staining. The myocyte apoptosis was measured with TUNEL staining. The protein expressions of AKT, phospho-AKT (P-AKT), Survivin, and phospho-BAD (P-BAD) were measured by Western blot.ResultsThe size of CMB was uniformly. The zeta potential of CMB was significantly higher than that of DMB (t=28.680, P=0.000). The CMB bound more plasmid DNA than the DMB (P<0.05). The luciferase activity of myocardium were higher in CMB group than in DMB group bothin vitro and in vivo measurements (P<0.05). There was no significant difference between groups in the ratio of signal intensity in anterior wall to posterior wall, ejection fraction (EF), and fractional shortening (FS) at 5 days after I/R (P>0.05), but the above indexes were significant higher in CMB and DMB groups than in control group at 21 days after I/R (P<0.05). Besides, the above indexes were significant higher in CMB group than in DMB group at 21 days after I/R (P<0.05). The infarct size was the smallest and infarct thickness was the thickest in the CMB group, followed by DMB group, control group at 21 days after I/R. The capillary and arteriolar densities of CMB and DMB groups were significant higher than those of control group at 21 days after I/R (P<0.05). Besides, the capillary and arteriolar densities of CMB group were significant higher than those of DMB group (P<0.05). The apoptotic cells were the most in the control group, followed by DMB group, CMB group at 3 days after gene transfection, showing significant differences between groups (P<0.05). The protein expressions of AKT, P-AKT, Survivin, and P-BAD were significant higher in CMB and DMB groups than those in control group at 3 days after gene transfection (P<0.05). Besides, these protein expressions were significant higher in CMB group than those in DMB group (P<0.05).ConclusionThe DNA-carrying capacity and gene transfection efficiency are elevated by CMB, although its physicochemical property is the same as DMB. When ultrasound-targeted AKT gene transfection is used to treat myocardial I/R injury in rats, delivery of AKT with the CMB can result in higher transfection efficiency and greater cardiac functional improvements compared to the DMB.
ObjectiveCD44 and CD54 are two specific biomarkers of gastric cancer stem cells and were used as targets in this study. The number of CD45–CD44+CD54+ cell subsets in peripheral blood of gastric cancer patients was detected by flow cytometry. Further, we combined these results with the clinicopathological characteristics of gastric cancer patients to analyze the significance of CD45–CD44+CD54+ cell subsets.MethodsFrom December 2016 to September 2017, 38 patients with gastric cancer in gastrointestinal surgery of West China Hospital of Sichuan University were included as the study object. The content of CD45–CD44+CD54+ cell subsets in their peripheral blood was detected by flow cytometry and its clinical significance was analyzed.ResultsThe median number of CD45–CD44+CD54+ cells were 541.9/mL (71.7–8 057.0/mL) in 38 patients and 555.9/mL (71.7–8 057.0/mL) in the group of patients with R0 resection. Patients without lymph node metastasis were found to have more CD45–CD44+CD54+ cells than patients with lymph node metastasis [941.4/mL (183.5–8 057.0)/mL vs 379.3/mL (71.7–2 269.7/mL, P=0.002], and more CD45–CD44+CD54+ cells in patients with TNM stage Ⅰ–Ⅱ than in TNM stage Ⅲ–Ⅳ [858.6/mL (183.5–8 057.0/mL) vs 364.6/mL (71.7–2 269.7/mL, P=0.015]. The patients with T3–4 stages (P= 0.025), N+ stage (P=0.009) and TNM Ⅲ–Ⅳ stage (P=0.012) had low ratios of the subgroup with high number of CD45–CD44+CD54+ cells, respectively. We made a more accurate judgment of N stage and TNM stage when we combined tumor size and the number of CD45–CD44+CD54+ cells together. However, there was no significant correlation between the number of CD45–CD44+CD54+ cells and other clinicopathological features and prognosis.ConclusionsThe number of CD45–CD44+CD54+ cell subsets is correlated with tumor progression, which might be used to predict TNM stage and N stage. However, the number of patients included in this study is too small, and the clinical significance of CD45–CD44+CD54+ subsets in gastric cancer patients needs to be further demonstrated by expanding the sample size.