| Objective:Establish the bilateral cecal and abdominal wall injury model to investigate the anti-adhesion effect of medical sodium hyaluronate gel(SHA).Calculate the ultimate load ratio and stiffness ratio of the muscle at the abdominal wall injury to the normal side muscle to see the healing of damaged muscle tissue.Investigate the acute toxicity of SHA in mice by i.p.Teste the effects of SHA on bacterial growth,spread of antibiotics,and tumor growth and metastasis to determine whether SHA could be used for postoperative anti-adhesion in patients with clinical abdominal or pelvic tumor resection.Method:(1)The experiment was randomly divided into sham operation group,model group,low dose group(0.15 m L/cm~2),high dose group(0.30 m L/cm~2)and commercially available control group(0.15 m L/cm~2).A needle-like bleeding wound of 1 cm×2 cm in the cecum near the side of the abdominal wall was operated and at the same time,a 1 cm×2 cm superficial muscle injury at the corresponding abdominal wall of the cecum was also created.The cecum was exposed to the air for 8 min.Then saline and different doses of SHA were applyed to the cecum injury and abdominal wall injury according to different groups.The cecum wound was placed directly opposite the abdominal wall wound back into the abdominal cavity.The abdominal wall wound was sewed layer by layer.30 days later the rats were killed by anesthetic.The abdomen was opened as"U"type and the abdominal adhesion was observed.The 5-grade classification method was used to evaluate the abdominal adhesions.A piece of 1 cm×2 cm muscle was cut from the injured side and the normal side.The tension instrument was used to measure the ultimate load of the muscles on both sides of the abdominal wall and calculate the stiffness ratio to investigate the effect of SHA on the healing of injured muscle tissue.(2)The growth curves of Escherichia coli(E.coli)and Staphylococcus aureus(SA)were determined to incubated with different concentrations of SHA.The effect of SHA on the proliferation of the two bacteria were observed respectively.The inhibition zone was determined by punch-diffusion assay with several antibiotics including cefoxime sodium,levofloxacin,gentamicin Sulfate,penicillin sodium and azithromycin,which were spotted alone or with the combination of SHA against E.coli and SA.The effect of the SHA on the efficacy of antibiotics was then observed in vitro.The effect of SHA on bacterial proliferation was investigated in mice by measuring the bacterial load in the peritoneal fluid.The effect of SHA on abdominal abscess was investigated in bacterial infected rats after cecum injury.(3)The effects of different concentrations of SHA on the growth and the migration of tumor cells were investigated by MTT assay and transwell assay in vitro in three tumor cells including Hela,CT26 and HCT116.An orthotopic transplantation model of colonic tumorin nude mice was established.The effect on the proliferation of cell HCT116 was investigated by comparing the tumor volume and tumor mass 4 weeks after inoculation.The effect on the metastasis of cell CT26 was investigated by comparing the tumor metastasis rate and the number of metastatic lesions of lung and liver in nude mice among the different experimental groups 3 weeks after inoculation.(4)To investigate the acute toxicity of SHA,different doses of SHA were injected in mice by i.p.The changes of body weight and clinical manifestations of the mice were observed in the following 14 days.The pathological changes of the organs of the mice was observed after 14 days.Results:(1)The adhesion scores of the model group and the sham operation group were 3.60±1.26 and 0.10±0.32(P<0.001),suggested that the adhesion models were successfully established.The adhesion score of the low dose of SHA group was 1.80±1.75,(vs model group,P<0.01).The high-dose of SHA group had an adhesion score of1.73±1.69,(vs model group,P<0.01).The limit load ratio and the stiffness ratio of the low-dose group or the high-dose group were not significantly different from the sham operation group or the model group respectively(P>0.05).(2)Different concentrations of SHA did not promote the growth of E.coli and SA.There was no statistically significance in the area of the inhibition zone between the antibiotics combined with SHA group and the antibiotics alone group(P>0.05).There was no significant difference in the bacterial load of the peritoneal fluid between the mice in the SHA group and the model group.In the experiment of the effect of SHA on the in vivo diffusion of levofloxacin and penicillin sodium,there was no significant difference in the abscess score of the abdominal cavity between the SHA group and the model group.The abscess scores of the antibiotics group and the antibiotics combination with SHA group were lower than the model group,and the differences were statistically significant.There was no significant difference in the abdominal abscess score between the antibiotics combined SHA group and the antibiotics group.(3)SHA did not promote the growth and metastasis of Hela,CT26 and HCT116 cells in vitro at different concentrations.Actually,SHA exhibited a certain inhibitory activity at higher concentration.In the orthotopic transplantation model of colonic tumor-HCT116,SHA did not promote the growth of cell HCT116.In the orthotopic transplantation model of colonic tumor-CT26,SHA inhibited CT26 tumor metastasis.(4)The weight and clinical manifestations of the mice after intraperitoneal injection of SHA were normal in the acute toxicity experiment.Conclusion:SHA has a significant anti-adhesion effect on postoperative abdominal adhesion in rats,without affecting wound healing.High concentration of SHA has an inhibitory effect on bacteria,and no effect on the exertion of antibiotics has been observed.SHA does not promote the growth and spread of tumor cells Hela,CT26 and HCT116;high concentrations of SHA in vitro inhibits the growth of tumor cells;SHA inhibits tumor cell metastasis both in vivo and in vitro.SHA has no acute toxicity to mice at dose of 500mg/kg.SHA can be used for postoperative adhesion prevention in patients with abdominal and pelvic tumors. |
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