| Objective:1) To evaluate the efficacy and safety of transarterial chemoemolization (TACE) combined with Radiotherapy (RT) or TACE monotherapy in the treamen of hepatocelluar carcinoma (HCC); 2) To analyze and initiate the potential mechanisms of radiation induced liver damage (RILD) and potential targets for treatment of RILD, find the key targets of the related signaling pathways. Methods:1) Patients with recurrent HCC who received the treatment of TACE puls RT (25 cases) or TACE monotherapy (50 cases) in our single center during Jan.2012 to June.2013 were retrospectively analyzed; 2) In vivo experiments:Forty healthy male Sprague-Dawley rats were randomly divided into model group (30 rats) and control group (10 rats). The right-half of rat liver in model group was irradiated with a single dose of 30Gy to establish radiation-induced hepatic injury model. The control group was false radiation in the same situation. Serum and liver tissues from the both group were collected at different time points (3 days,5 days and 10days) after irradiation. Changes of rat serum AST, ALT, ALP, TB, DB were detected with automatic biochemical analyzer Pathomorphological change in the liver was observed with hematoxylin-eosin and Masson staining by optical microscope. The protein expression of TGF-β, α-SMA and p-Akt (S473) was assayed by immunohistochemistry. Weight measurements and blood was obtained from the rats and liver tissues were collected for histological and apoptotic analysis; 3) HSC-T6 were exposed to the different doses of 6 MV X-ray (OGy, lOGy,20Gy) irradiation and PI3K/Akt signaling pathway inhibitor LY294002. The apoptosis rate was detected by Flow Cytometry (FCM). The cell supernatants expression of transforming growth facter β1 (TGF-β1) was detected by ELISA assay.The expression of Akt and α-smooth muscle actin (a-SMA) mRNA were detected by real-time PCR. The expression of p-Akt was detected by Western blot assay. Results:1) Follow-up of 75 patients with HCC, according to the treatment methods are divided into joint group and conventional TACE alone.Simple set of 50 cases with conventional, median survival time was 7 months; Joint group of 25 patients, median survival time was 13 months, draw Kaplan Meier survival curve and the log-rank test shows that joint group compared with pure group with conventional time difference is statistically significant (chi-square=28.72, P<0.05. Combined search Wu Dehua, etc, combined with conventional and local radiotherapy in the treatment of 94 cases of surgery in patients with primary liver cancer, with 48～60Gy,8～10 times,21～24 d completed,3 months after treatment completely relieve 12 cases, partial in 73 cases,6 cases stability, progress in 3 cases.1,2,3 year survival rate is respectively 93.6%and 53.8%,26.0%, the median surial 25.0 a month; 2) The expression of AST, ALT, ALP, TB, DB were significantly higher in the model group than that in the control group at different time points (3 days,5 days and 10days) after irradiation. HE and Masson staining showed that the expression of collagen fibers is higher in the surrounding liver blood sinus in the radiation induced acute liver injury in rats. The expression of TGF-beta 1, alpha-SMA, p-Akt (S473) gradually increased with irradiation was observed after prolonged by immunohistochemical method of time, the difference was statistically significant (P< 0.000) by IPP analysis; 3) FCM showed that, compared with the control group, the apoptosis rate of the 10 and 20Gy group increased with the increase of irradiation dose (t=8.43,11.63, P<0.05); The apoptosis rate decreased after the same irradiation dose plus inhibitor group (t=8.09,4.89, P<0.05). ELISA assay showed that, compared with the control group, the expression of TGF-β1 increased with the increase of irradiation dose in the 10 and 20Gy group (t=11.51,19.02, P<0.05); Compared with the simple irradiation group, the expression of TGF-β1 decreased after the same irradiation dose plus inhibitor group (t=6.17,10.04, P<0.05). RT-PCR assay showed that, compared with the lOGy group, the expression of Akt and a-SMA mRNA increased in the 20Gy group (t=4.89,7.80, P<0.05), it decreased in the 10Gy+LY294002 group (t=13.66,15.11, P<0.05); Compared with the 20Gy group, the expression of Akt and a-SMA mRNA decreased in the 20Gy+LY294002 group (t=6.20,6.85, P<0.05). Western blot assay showed that, compared with the control group, the expression of p-Akt increased with the increase of irradiation dose in the 10 and 20Gy group (t=5.74,31.23, P<0.05); Compared with the simple irradiation group, the expression of p-Akt decreased after the same irradiation dose plus inhibitor group (t=10.34,23.84, P<0.05). Conclusions:1) TACE combine with RT is superior to TACE monotherapy with respect to the overall survival and tumor-free survival of HCC patients.The differentiation degree and recurrence interval of carcinoma in correlated with the prognosis of patients; 2) In a single large dose of radiation processes lead to the occurrence of acute radiation injury of liver, accompanied activation of hepatic stellate cells, increased secretion of TGF-beta 1, activation of PI3K/Akt signal pathway. PI3K/Akt signaling pathway may become a potential therapeutic target for radiation-induced liver injury. |
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