本文選題：兔VX2肢體軟組織腫瘤模型 + 容積弧形調(diào)強放療　； 參考：《新疆醫(yī)科大學》2015年博士論文

【摘要】：目的:1)通過超聲引導建立兔VX2肢體軟組織腫瘤模型,為后期的干預研究和影像學評價奠定基礎;2)在腫瘤模型上實施新輔助放療,應用容積弧形調(diào)強放療-同步推量(VMAT-SIB)技術在腫瘤毗鄰骨骼的區(qū)域提高照射劑量,局部產(chǎn)生放射性損傷,制作一個壞死帶定義為外科邊界,評價制作外科邊界的可行性、有效性;3)在腫瘤模型上,實施大分割放療制作外科邊界,評價其安全性。方法:1)選用雄性新西蘭兔30只,隨機分為對照組6只(在超聲引導下注射生理鹽水0.5ml),模型組24只(在超聲引導下注射腫瘤組織混懸液0.5ml)。兩組應用磁共振彌散成像(MRDWI)和三維超聲觀察腫瘤的形態(tài)、信號特點及體積變化。對照組在采集完影像后處死動物觀察組織學變化。模型組于第14天、21天、28天隨機分批處死6只動物觀察組織學變化。剩下6只動物不處死,觀察帶瘤生存時間;2)選用30只腫瘤模型兔,隨機分為對照組:10只;調(diào)強放療(IMRT)組:10只;VMAT-SIB組:10只。三組在放療后一周進行影像學采集:MRDWI、三維能量多普勒超聲血管成像(3D-PDA)。之后,處死動物,觀察組織學變化以及采用免疫組化檢查腫瘤組織的增殖、凋亡、血管密變化;3)選用40只腫瘤模型兔,隨機分為對照組:10只;VMAT-SIB3Gy組:10只;VMAT-SIB4Gy組:10只;VMAT-SIB5Gy組:10只。四組在放療后,參照CTCAE分級觀察皮膚的大體情況。采用HE染色觀察皮膚和骨組織的形態(tài)學改變。采用透射電鏡觀察骨細胞超微結(jié)構(gòu)改變。采用熒光原位雜交(FISH)測定皮膚組織PDGF-B、EGF的表達情況。采用實時熒光定量PCR測定皮膚組織VEGF、bFGF的基因表達量。結(jié)果:1)三維超聲和MRDWI證實,24只實驗動物的種植成瘤率達100%,24只腫瘤模型兔全部存活。三維超聲結(jié)果,對照組在注射部位未見異常改變。模型組隨著時間的增加腫瘤體積逐漸增大,與對照組比較其差異具有統(tǒng)計學意義(P0.05)。MRDWI顯示:對照組未見明顯的異常信號。模型組在左側(cè)后肢可見腫瘤影像,腫瘤組織在DWI圖呈高信號;在ADC圖呈低信號。組織學結(jié)果,對照組:HE可見正常的橫紋肌及支氣管肺泡組織。模型組:第14天取材,HE可見生長旺盛的瘤細胞呈不規(guī)則致密排列,侵犯橫紋肌。第21天取材,HE可見腫瘤區(qū)與片狀壞死區(qū)邊界不清,腫瘤生長區(qū)內(nèi)細胞較密集。第28天取材,HE可見部分的腫瘤細胞核固縮、細胞核碎裂。第60天取材,HE可見腫瘤邊緣仍有腫瘤細胞,腫瘤實質(zhì)可見壞死細胞,伴有炎細胞浸潤及纖維組織增生。解剖胸腔,見兩肺彌漫性轉(zhuǎn)移結(jié)節(jié),部分小結(jié)節(jié)相互融合,HE可見腫瘤細胞侵犯支氣管及肺泡;2)在放療過程中,IMRT組有2只腫瘤模型兔死亡,一只動物因麻醉過量致死,一只動物由于腫瘤破潰至皮膚給予安樂死。三組之間的感興趣區(qū)(毗鄰骨骼的腫瘤組織)在DWI圖和ADC圖均表現(xiàn)出顯著的不同。對照組在DWI圖呈高信號;在ADC圖呈明顯低信號。IMRT組在DWI圖呈欠均勻低信號;在ADC圖呈高信號。VMAT組在DWI圖呈低信號;在ADC圖呈高信號。3D-PDA觀察:三組之間的VI、FI及VFI均表現(xiàn)出顯著的不同,IMRT組和VMAT組明顯低于對照組。組織學結(jié)果,對照組:HE可見腫瘤細胞侵犯橫紋肌,正常的細胞結(jié)構(gòu)受到損害。IMRT組:HE可見腫瘤實質(zhì)及腫瘤邊界組織大片的壞死細胞,伴有炎細胞浸潤及纖維組織增生。VMAT組在腫瘤毗鄰骨骼的區(qū)域可見一壞死帶形成,鏡下見細胞壞死明顯,纖維組織增生,周圍有炎細胞浸潤。放療后,IMRT組和VMAT組的增殖活性降低、血管密度降低、凋亡增加;3)對照組、3Gy組和4Gy組的動物在放療期間和放療后,局部皮膚未見異常。5Gy組有8只動物在放療期間,局部皮膚出現(xiàn)了輕度紅斑和干燥性脫屑。對照組、3Gy組和4Gy組在鏡下可見正常的皮膚組織,皮膚表皮各層結(jié)構(gòu)完整。5Gy組可見表皮有大量的角化物質(zhì),上皮下有炎細胞浸潤。對照組、3Gy組和4Gy組在鏡下可見骨細胞形態(tài)正常。5Gy組在鏡下可見骨細胞排列松散,細胞表現(xiàn)出明顯的萎縮。透射電鏡結(jié)果顯示,對照組、3Gy組和4Gy組的骨細胞形態(tài)正常。5Gy組可見粗面內(nèi)質(zhì)網(wǎng)擴張,呈囊泡狀樣改變。FISH:EGF和PDGF-B的mRNA結(jié)果:對照組與3Gy組、4Gy組、5Gy組比較,其差異具有統(tǒng)計學意義(P0.05)。實時熒光定量PCR:VEGF mRNA結(jié)果:對照組與3Gy組、4Gy組比較,其差異具有統(tǒng)計學意義(P0.05),但與5Gy組比較未見明顯差異(P0.05)。bFGF mRNA結(jié)果:對照組與3Gy組、4Gy組比較其差異具有統(tǒng)計學意義(P0.05),但與5Gy組比較未見明顯差異(P0.05)。結(jié)論:1)通過超聲引導注射腫瘤組織懸液,成瘤率高,腫瘤生長范圍較局限。影像學和組織學結(jié)果提示:后期的干預研究應在種植腫瘤后的第14天左右進行,以降低腫瘤內(nèi)部液化壞死對實驗結(jié)果的影響;2)應用VMAT-SIB技術在腫瘤模型上實施新輔助放療具有一定的可行性。MRDWI、3D-PDA、組織學證實,在腫瘤毗鄰骨骼的區(qū)域制作的壞死帶,能夠有效的定義為外科邊界;3)在腫瘤模型上實施大劑量分割放療,結(jié)果提示VMAT-SIB 3Gy組和VMAT-SIB4Gy組的動物能夠耐受早期的局部毒性反應,是一個相對較安全的劑量分割方案。
[Abstract]:Objective: 1) the tumor model of VX2 limb soft tissue in rabbits was established by ultrasonic guidance to lay the foundation for the later intervention study and imaging evaluation; 2) to implement new adjuvant radiotherapy on the tumor model, and to improve the radiation dose in the region adjacent to the adjacent bone with volume arc intensity modulated radiotherapy synchronous push (VMAT-SIB) technology, and to produce radioactivity damage locally. A necrotic band was defined as the surgical boundary, the feasibility and effectiveness of making the surgical boundary was evaluated. 3) in the tumor model, the surgical boundary was made by large fractionation radiotherapy, and the safety was evaluated. Method: 30 male New Zealand rabbits were selected to be randomly divided into 6 control groups (0.5ml under ultrasound guidance), and 24 in the model group. Ultrasound guided injection of the tumor tissue suspension 0.5ml. The two groups were treated with magnetic resonance diffusion imaging (MRDWI) and three-dimensional ultrasound to observe the tumor morphology, signal characteristics and volume changes. The control group died of animal observation after the collection of images. The model group was executed at fourteenth days, 21 days, 28 days and 6 animals were killed at random to observe histological changes. The remaining 6 animals were not killed, and the survival time of the tumor was observed; 2) 30 rabbit models were selected and randomly divided into the control group: 10; the intensity modulated radiotherapy (IMRT) group: 10; the VMAT-SIB group: 10. The three groups were collected for one week after radiotherapy: MRDWI, and three-dimensional energy Doppler ultrasound angiography (3D-PDA). Then, the animals were executed and histological changes were observed. And using immunohistochemistry to examine the proliferation, apoptosis, vascular density change of tumor tissue; 3) selected 40 tumor model rabbits, randomly divided into control group: 10 rats, group VMAT-SIB3Gy, 10; group VMAT-SIB4Gy, 10; group VMAT-SIB5Gy: 10. After radiotherapy, the general condition of skin was observed by CTCAE classification. HE staining was used to observe the skin and bone tissue. Morphological changes. The ultrastructural changes of the bone cells were observed by transmission electron microscopy. The expression of PDGF-B and EGF in skin tissue was measured by fluorescence in situ hybridization (FISH). The expression of VEGF and bFGF in skin tissue was measured by real-time fluorescence quantitative PCR. Results: 1) three dimensional ultrasound and MRDWI confirmed that the growth rate of 24 experimental animals was 100%, 24 The tumor model rabbits were all alive. The results of three-dimensional ultrasound showed no abnormal change at the injection site. The tumor volume in the model group increased gradually with the increase of time. Compared with the control group, the difference was statistically significant (P0.05).MRDWI showed that there was no obvious abnormal signal in the control group. The model group showed tumor image and tumor on the left hind limb. Tissue showed high signal on DWI map, low signal on ADC map. Histological results, control group: normal rhabdomyoma and bronchioloalveolar tissue were seen in HE. Model group: fourteenth days of extraction, HE visible growth of tumor cells were irregular and dense arrangement, invasion of rhabdomyoma. Twenty-first days, HE visible tumor area and flaky necrotic area boundary was unclear, tumor birth The cells in the long area were denser. After twenty-eighth days, the nuclei of the tumor were recovered and the nuclei were broken. The tumor cells were found on the edge of the tumor on the sixtieth day, and the tumor cells were still found on the edge of the tumor. The tumor was visible with necrotic cells, with inflammatory cells infiltration and fibrous tissue proliferation. The thoracic cavity was dissected, two pulmonary diffuse nodules were seen, and some small nodules fused, HE It can be seen that tumor cells invade bronchi and alveoli; 2) during the course of radiotherapy, 2 tumor model rabbits in group IMRT died, one animal died of overdose of anesthesia, and one animal broke into the skin for euthanasia due to the collapse of the tumor to the skin. The region of interest between the three groups (adjacent to the bone tumor group) showed significant differences between the DWI map and the ADC map. The group in the DWI map showed high signal, the ADC graph showed the low signal.IMRT group in the DWI diagram and the low signal in the DWI diagram; the high signal group in the ADC diagram showed the low signal in the DWI diagram; the VI, FI and VFI were significantly different between the three groups, and the group and the group were obviously lower than the control group. Histology results, control group: control group: It can be seen that the tumor cells infringe the rhabdomyosus muscle, and the normal cell structure is damaged in the.IMRT group: HE can be seen in the tumor parenchyma and the necrotic cells of the tumor boundary tissue, with the infiltration of inflammatory cells and the proliferation of fibrous tissue in the.VMAT group, a necrotic zone is visible in the area adjacent to the adjacent bone, and the necrosis of the cells, the proliferation of fibrous tissue, and the surrounding area are observed under the microscope. After radiotherapy, the proliferation activity of IMRT group and VMAT group decreased, blood vessel density decreased, apoptosis increased; 3) control group, 8 animals in group 3Gy and 4Gy group had no abnormal.5Gy group during radiotherapy and radiotherapy, and the local skin appeared mild erythema and dryness desquamation during radiotherapy. The control group, 3Gy group and 4Gy were in the control group. The normal skin tissue was seen under the microscope, and the epidermis of all layers of the skin had a large number of keratinized substances and infiltration of inflammatory cells under the epithelium. In the control group, group 3Gy and group 4Gy showed that the bone cell morphology in the normal.5Gy group was visible under the microscope, and the cells were loosely arranged and the cells showed obvious atrophy. The transmission electron microscope showed that the transmission electron microscope showed that the cells were obviously atrophied under the microscope. In the control group, the 3Gy group and the 4Gy group, the normal.5Gy group showed the dilation of the rough endoplasmic reticulum and the mRNA results of the vesicular like change of.FISH:EGF and PDGF-B. The control group was statistically significant compared with the 3Gy group, 4Gy group and 5Gy group (P0.05). The real time fluorescence quantitative PCR:VEGF mRNA results were compared with those in the control group. Statistical significance (P0.05), but compared with the 5Gy group, there was no significant difference (P0.05).BFGF mRNA results: in the control group and the 3Gy group, the difference in the 4Gy group was statistically significant (P0.05), but there was no significant difference between the group and the 5Gy group (P0.05). Conclusion: 1) the tumor growth rate was high and the growth range of the tumor was limited by ultrasound guided injection of tumor tissue suspension. The results of image and histology suggest that the later intervention study should be carried out about fourteenth days after the tumor is planted to reduce the effect of internal liquefaction and necrosis of the tumor to the experimental results. 2) the application of VMAT-SIB technology to the neoadjuvant radiotherapy on the tumor model has a certain feasibility,.MRDWI, 3D-PDA, histologically confirmed, in the region adjacent to the bone. The necrotic zone made by the domain can be effectively defined as the surgical boundary; 3) the large dose fractionation radiotherapy on the tumor model indicates that the animals in the VMAT-SIB 3Gy group and the VMAT-SIB4Gy group can tolerate the early local toxicity, which is a relatively safe dose division scheme.
【學位授予單位】：新疆醫(yī)科大學
【學位級別】：博士
【學位授予年份】：2015
【分類號】：R738.6;R-332

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