| Blackground and ObjectivesThere are more than 1000 kinds of bacteria in human intestine,the total number of which is close to 1012-1014,that is gut microbial.It spreads all over the oral cavity,intestines,skin,genitourinary system,respiratory system and so on.It communicates with the host in many ways and affects its host.Antibiotics are widely used in the treatment of various infectious diseases,especially in surgery,antibiotics are often used prophylactically before surgery,and large doses of antibiotics are used to treat postoperative infection.At present,a large number of studies have confirmed that this change has an impact on bone mass and bone microstructure.However,little is known about the effect of intestinal flora changes on fracture healing.In this paper,we discussed the effect of antibiotic induced intestinal flora damage on bone healing process in two cases of chronic osteomyelitis(CO)after internal fixation alone and infection after internal fixation.MethodsC57BL/6 mice was divided into three groups,including the antibiotic-treated chronic osteomyelitis(Ab-CO,N=28),the CO model group(CO,N=28),the antibiotic-treated implant(Ab-implant,N=15),the implant group(Implant,N=15),and the blank group(Blank,N=15).Mice in the Ab-CO group and Ab-implant groups were additionally treated with an antibiotic cocktail from day-7 to day 28 to explore the role of the gut microbiota on the healing process of fractured and/or infected bone.An antibiotic cocktail was prepared as previously described containing kanamycin(0.8 mg/mL;Sigma),gentamicin(0.07 mg/mL;Sigma),colistin(0.1135 mg/mL;Sigma),metronidazole(0.43 mg/mL;Sigma)and vancomycin(0.09 mg/mL;Sigma).The group of CO,Implant and Blank began to drink distilled water at D-7 days,until D28 days.On D 0 day,Staphylococcus aureus was used to model bone infection in Ab-CO and CO;Ab Implant and Implant were used to model bone fracture.Mice in the blank group were reared from beginning to end without any treatment.Body weight and serum levels of tumor necrosis factor-αand interleukin-6 were monitored.Gut microbiota composition was assessed by 16S rRNA sequencing and bone condition was analyzed by micro-CT and staining with hematoxylin and eosin,Safranin O-fast green and Goldner’s trichrome.Osteoblastogenesis and osteoclastogenesis were assessed by detection of tartrate resistant acid phosphatase and osterix expression.ResultMice in Ab-CO and Ab-Implant grew slowly with a treatment of antibiotic cocktail from day-7 to day 0,and mice have strated dying since day 3.Significant differences in survival rates were observed between the CO and Ab-CO groups at day 28(P=0.0471).The intestinal mucosa of mice taking antibiotics was incomplete and even exfoliated.Perturbations of the gut microbiota led to delayed fracture union along with increased levels of pro-inflammatory cytokines in serum(p<0.01),lower bone mass(p<0.05),deficient endochondral ossification and bone formation,reduced osteoblastogenesis(p=0.026)and enhanced osteoclastogenesis(p=0.017).We detected no differences in cytokines levels,bone mineral density or bone formation,potentially due to extreme damage to bone structure.In summary,we showed that antibiotic treatment perturbs gut microbiota and significantly interferes with bone fracture union by increasing circulating pro-inflammatory cytokine levels,inhibiting osteoblastogenesis and enhancing osteoclastogenesis,and leading to lower bone mass and higher mortality following infection.Ours is the first report to elucidate how antibiotic-induced alterations of the gut microbiota affect bone fracture union.These data advance our understanding of the role of gut microbiota disorders in osteoblastogenesis and osteoclastogenesis following bone fractures with or without infection.Conclusion1.High-dose antibiotic treatment induces significant gut microbiota dysregulation in mice2.Perturbation of gut microbiota impaired host resistance to CO and enhanced the pro-inflammatory response3.Antibiotic dysregulation of gut microbiota reduced bone mineral density(BMD)4.Gut microbiota dysregulation induced by antibiotics interfered with the process of bone formation5.Gut microbiota dysregulation induced by antibiotics enhanced osteoclast activity and inhibited osteoblast function... |
