The Study Of The Effect Of Machine Perfusion Combined With Antibiotics Prevents Donor-Derived Infection In The Rat Model

Objective KT(Kidney transplantation,肾移植)is the therapy of choice for end-stage kidney failure.However,the growing disparity between the supply of organs and number of recipients awaiting transplant has led to the acceptance of many marginal donors,such as those with a high risk of transmitting pathogens to recipients.Transplantation of active infected donor organs into recipients will increase the risk of DDI(Donor-derived infection,供体来源性感染).MP(Machine perfusion,机械灌注)is an emerging preservation alternative to SCS(Static cold storage,静态冷保存)and it is an essential platform of organ preservation,quality assessment,and injury repair.The prophylactic treatment and the utilization of donors with a high risk of DDI caused great controversy.Because the models for further research are lacking,the possible mechanism,the timing and effectiveness of preventive treatment are still unclear.Therefore,we established the bacteremia model in rats,rat kidney MP and KT models to systematically investigate the possible mechanism and effectiveness of preventing DDI under ideal conditions combined with AB(Antibiotic,抗生素).Methods(1)We established a rat model of bacteremia,inoculated with E.coli(Esche richia coli,大肠杆菌)or CRKP(Carbapenem-resistant Klebsiella pneumoniae,耐碳青霉烯类肺炎克雷伯杆菌)to investigate bacterial load of organs and blood in differ ent time point following inoculation.(2)Donor kidneys were procured after inoculatio n with bacteria for 24 hours.The kidneys were PFAB(Preflushed with antibiotic,抗生素溶液预冲洗)before they were procured or treated with NMP(Normothermic ma chine perfusion,常温机械灌注)、HMP(Hypothermic machine perfusion,低温机械灌注)and SCS combined with antibiotic for 3 hours.The effect of eliminating bacteri a before and after treatment was detected using immunofluorescence and quantitative bacterial tissue cultures.The NMP perfusate was collected to test the level of exogeno us creatinine to evaluate the kidney function.(3)According to the above methods,a ra t model of drug-resistant bacteremia was established,the kidneys were transplanted af ter ex vivo treatment to evaluate the effectiveness and safety of different preservation methods combined with antibiotic by in vivo research.(4)The status of recipient rats was monitored,when they reached humane endpoint,the kidneys,liver,and lungs wer e procured and cultured.Individual colonies of different organs were selected separate ly for validation of bacterial source using matrix-assisted laser desorption ionization-ti me of flight mass spectrometry,PCR(Polymerase chain reaction,聚合酶链反应),an d genetically sequenced.Results(1)In the rat model of E.coli inoculated via tail vein,the number of bacteria in blood reached the highest at 16 h,while the amount of bacteria in organs reached the peak at 24 h.Per unit weight,the bacterial load in different organs were arranged from high to low as liver,lungs and kidneys.The bacterial load in the lungs of CRKP infected rats was significantly higher than that of E.coli infected rats.(2)Before ex vivo treatment,bacteria were intensively distributed in the renal cortex,the number of bacteria in the kidneys showed no significant difference after treatment compared with before treatment in the SCS+AB,PFAB+SCS+AB and HMP groups.However,the renal bacterial population in the HMP+AB and NMP+AB groups were significantly reduced after treatment.Compared with the normal control and CRKP groups,there was no significant difference in exogenous creatinine clearance in the CRKP+AB group,indicating that transit high-dose antibiotic treatment did not disturb renal function.(3)The rats in the SCS+AB group after transplantation exhibited widespread and serious infection of the abdominal cavity and organs,anastomotic bleeding occurred in one case.However,the rats in the HMP+AB group were in good condition.(4)The BLAST analysis of the bacterial sequence of recipient rats indicated that the tested bacterial sequence was consistent with that of the standard strain and confirmed transmission of bacterial infection from donors to recipients.Conclusions This study used the model of donor kidney infected caused by bacteremia and KT to systematically verify the feasibility and efficacy of ex vivo renal MP combined with antibiotic in treating donor organ infection.It’s a good way to prevent DDI,expand the donor pool and improve the safety of KT.