| Objective: The change of mouse diaphragma after treated with cisplatincombined with17-AAG observed by HE staining, SEM and TEMtechnique.In order to provide information for the biology of tumor therapy.Methods: Select50BALB/c mouse, SPF, weight (20~21) g, Standardparticle feed, Free feeding and drinking water. The mice were randomlydivided into5groups:①Blank control group②Model group③17-AAGgroup④cisplatin group⑤cisplatin combined with17-AAG group.Each grouphas10mice.The blank control group do not treated, and the remaining fourgroups of intraperitoneal injection of H22tumor cells resulting in tumormodels.The5groups of mice were given different drug intervention. All themice were killed after10days.1.Observe the general condition of the mice andMeasure the weight of mice.2.Samples of diaphragmas were obtained afterbeing infused with4%polyformaldelryde, After different methods oftreatment.3.With2N NaoH at room temperature for2hours,2N NaoHdigestion digestion for6days at room temperature, ultrasonic treat1min,routine dehydration, drying, coating.Using scanning electron microscopicobserve the structure changes of mice diaphragm myoepithelial andsubepithelial connective tissue.4.Cut off a part of diaphragm,Observed bytransmission electron microscope.Results:1.General condition of the mice Modelld ten days later model group and17-AAG group's ascites increased significantly. The entire abdomen swellssignificantly and prominent spherical,Weight increased significantly.All themouse fatigue, autonomic activity decreased, less fur and lose luster. Mouse ofcisplatin group have a good growth state, normal hair, without ascites, and state of mind is good. Compared with the mice of blank control group thereare no significant differences between them.17-AAG combined with cisplatinon mice status were between model mice and cisplatin mice. Weight thanbefore to increase. But obviously not as good as the model group and17AAGgroup.2.HE staining results:Blank control group of peritoneal mesothelial cellsclose to the lower muscle cells and the subepithelial connective tissue is notobvious. Cells were flat,tightly packed. The nucleus is a flat oval, located atthe cell center. Peritoneal mesothelial cells of the model group increased,rounded, cubic shape. Nuclear enlargement and turn round. At the same timewe can see that the epithelial surface of tumor cell invasion and subepithelialconnective tissue proliferation.The number of blood vessels, lymphatic vesselsincreased. Part of the tumor cells through the gap between the epithelial cellsinto the connective tissue lumen, infiltration into the muscle cell surfacethrough the lumen.17-AAG group and Model group have Similarresults.Cisplatin group of peritoneal mesothelial cells were flat,tightly packed.The nucleus is a flat oval, located at the cell center. There are a fewhyperplasia of subepithelial connective tissue, but no tumor cell invasion.Peritoneal mesothelial cells of cisplatin combined with17-AAG grouparranged in rules. Cell gap compared with the normal group increases. Cellsincrease, rounded, cubic shape, nuclear enlargement and turn round.There aretumor cell invasion epithelial surface.Subepithelial connective tissueproliferation. Only see the individual tumor cells into the connective tissuelumen through the epithelial cell gap. No muscle cells on tumor cellattachment.3.The SEM results: Cell surface were observed by scanning electronmicroscope: In the blank control group,the surface of squamous cells ofphrenic myoepithelial cells have short microvilli. Cell borders are visible,adjacent cells are closely connected, there is no peritoneal poredistribution.The surface microvilli cubic cell were short and sparse.there arescattered or in clusters of the peritoneal stomata between them. Two kinds of cells were banded alternating distribution.The surface of phrenic myoepithelialcells is covered with a layer of tumor cells. Cubic cell morphological diversity,irregular arrangement.The surface microvilli of cubic cell increasedsignificantly.There are no scattered or clusters of the peritoneal stomatabetween the cells. Two kinds of the diaphragm epidermal cells were bandedalternating distribution and regularly arranged.The microvillous of cubic smallcell surface increased significantly,but the microvillous of the flat cell werereduction. Adjacent cells are connected closely, and the cell borders wereindistinct.In the17-AAG group,peritoneal mesothelial cells intermingled withlarge tumor cells. Adjacent cells are connected closely,and no peritoneal poredistribution.The microvilli of flat cell were reduce or loss. The cell boundaryis not clear.The microvilli of cubic cell surface was similar to the themicrovilli of cubic cell surface of the blank control group.There are noperitoneal hole between the cells.After2N NaoH solution at room temperaturedigest6days,epithelial fibrous connective tissue of the mice in each grouphave no significant difference. There are a lot of mc which mainly distributedin diaphragmatic muscular parts of the peritoneal subcutaneous.4.TEM results: Diaphragmatic peritoneal mesothelial cells of blankcontrol group was flat, closely arranged. The nuclei were flat oval, located atthe cell center. Peritoneal mesothelial cells of the model group increased,rounded, cubic shape. The subepithelial connective tissue hyperplasiaobviously. A large number of tumor cells through the epithelium into theconnective tissue lumen. Diaphragmatic peritoneal mesothelial cells ofcisplatin group was flat, closely arranged. The nucleus is a flat oval, located atthe cell center. The subepithelial connective tissue hyperplasia,and there aresome large tubular structure in it but no tumor cells in it. Peritonealmesothelial cells of the17-AAG group increased, rounded, cubic shape. Thesubepithelial connective tissue hyperplasia obviously. There are some largetubular structure and tumor cells in connective tissue.Cisplatin combined with17-AAG group and the blank control group have Similar results:Diaphragmatic peritoneal mesothelial cells was flat, closely arranged. Conclusions:1.The diaphragmatic epithelial cells of the mouse which peritoneal cavityhave malignant ascites turn round and cuboid.The Structural change ofepithelial cells rise the intercellular Space. There are a large number of tumorcells distributed between the epithelial cells.Tumor cells arrange close-up andcan not see peritoneum stoma. This showed that a large number of tumor cellsblock the lymphatic stomata and MC.The formation of ascites was related tothe blockage of lymphatic return and peritoneal fluid malabsorption.2.The form of diaphragmatic epithelium in cisplatin combined with17-AAG group is better than17AAG group but worse than cisplatin group. |
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