Experimental Study Of Novel Layered Double Hydroxides Heparin Nanocomposites On Improving Microcirculation Of Septic Rats

Background Sepsis is a serious life-threatening organ dysfunction caused by the body response disorders due to infection or suspected infection,among which microcirculation dysfunction is not only the pathophysiological manifestation of sepsis,but also one of the initiating factors of the occurrence and development of sepsis.Low molecular weight heparin(LMWH)is a commonly used anticoagulant in clinical practice,and previous studies have investigated LMWH adjuvant therapy for septic coagulopathy.However,it has some limitations.Layered double hydroxides(LDH)are an emergent class of inorganic lamellar nanomaterials having a 2D structure.Due to the interlayer anion exchangeability of LDH,some anionic drug molecules can be intercalated into LDH layers to achieve controlled release of drugs under physiological conditions.Previous studies have shown that negatively charged LMWH can be intercalated into LDH layers,and the conjugation of fibrin antibody onto the LDH surface can make it target to the site of thrombi,then the continuous release of LMWH from LDH is enough to minimize thrombi.However,there are few studies on targeted therapy of microthrombi in sepsis.Purpose to construct fibrin antibody-LMWH-LDH nanocomposites to deliver the antibody-targeted drug to the microthrombus,so as to examine the effect on improving the microcirculation of septic rats.Methods FITC-20LMWH-LDH was firstly synthesized by a hydrothermal synthesis method,and then a certain amount of bovine serum albumin(BSA)was added and fibrin antibody(Ab)was conjugated onto BSA,so that Ab-BSA/BSA was successfully adsorbed on the surface of LDH nanoparticles,and finally FITC-Ab-BSA-20LMWH-LDH nanocomposites were synthesized.It was characterized by Fourier transform infrared spectroscopy(FTIR),X-ray diffraction(XRD),transmission electron microscopy(TEM),particle size and Zeta potential,and the drug release characteristics under different p H values were determined.The thrombus targeting ability of the nanocomposite was demonstrated by capillary tubule studies in vitro and intestinal microcirculation investigations in vivo.In the drug evaluation experiment,thirty-eight healthy male SD rats were randomly divided into five groups: the Sham+NS group,the Sepsis+NS group,the Sepsis+LMWH group,the Sepsis+20LMWH-LDH group,and the Sepsis+Ab-BSA-20LMWH-LDH group,and the caecal ligation and perforation(CLP)method was used to establish sepsis models.Rats in five groups were intravenously injected separately with 1 m L normal saline(NS),1 m L NS,1 m L LMWH,1 m L20LMWH-LDH and 1 m L Ab-BSA-20LMWH-LDH at 8 hours after surgery.At 24 hours after surgery,we observed the intestinal microcirculation of rats using a side-stream dark field imaging(SDF).Then all rats were euthanized.Blood was obtained by cardiac puncture and the main tissues including heart,liver,lung,and kidney were harvested for further histological examination.Results The successful synthesis of FITC-Ab-BSA-20LMWH-LDH nanocomposites was proved by the characterization of FTIR,XRD,TEM,particle size and Zeta potential.The drug release pattern of the nanocomposites is a gradual and biphasic release pattern,that is,rapid early release followed by continuous slow release with the release rate much quicker in acidic environment.Capillary tubule studies and intestinal microcirculation investigations showed that the green-fluorescent nanocomposites accumulated around microthrombi,proving that nanocomposites have the ability of targeting microthrombi.The observation of intestinal microcirculation,blood test indexes and the histopathology of important organs 24 h after surgery showed that the Ab-BSA-20LMWH-LDH treatment group improved microcirculation to a greater extent,and alleviated inflammation and organ dysfunctions of heart,lung,liver and kidney in septic rats.Conclusion FITC-Ab-BSA-20LMWH-LDH nanocomposites was successfully synthesized by hydrothermal synthesis method.The nanocomposites have a gradual and biphasic drug release pattern with the release rate much quicker in acidic environment.Moreover,they have the ability of targeting microthrombi,which can maximize antithrombotic,reduce bleeding risk,improve microcirculation,alleviate inflammation and organ dysfunctions in septic rats.