| Lipopolysaccharide (LPS), a component of cell wall in gram-negative bacteria, can induce organism to develop the same pathologic processes with the infection of gram-negative bacteria as a pathogen endotoxin. In this study, female rabbits were injected with LPS to induce a reversible and non-symptoms infection of gram-negative bacteria under normal physiological condition, and our aim was to reveal the effects of gram-negative bacteria infection on expression level of Toll-like receptor2and4(TLR2and TLR4) and downstream inflammatory cytokines (IL-1β, TNF-α, IL-6) in different segments of uterus.LPS-injection dosage and the time-course for real-time PCR was determined according to the pre-experiment of the content of white blood cell (WBC). Namely,20rabbits were divided randomly into5groups (n=4), and five groups were injected with saline (control group), and with LPS at the dosage of0.5,1,2and4mg/kg body weight (WB) respectively. The venous blood was collected at Oh (before injection), and1.5,3,6,12and24h of postinjection for measuring the content of WBC. And the results showed that the change of WBC content in blood has no dose-dependent effects at the dosage of0.5to4mg/kg WB by1.5to6h of LPS postinjection, but the content of WBC in all LPS-injection groups were significantly lower than control group (P<0.05). The content of WBC in0.5,1and2mg/kg WB LPS-injection groups recovered to the level in control group by12h of postinjection, but it in4mg/kg WB LPS-injection group recovered the level in control group by24h of postinjection. However, the dosage of1,2and4mg/kg WB LPS-injection caused rabbits dead for12or24h after injection. So we confirmed that0.5mg/kg WB was as an appropriate LPS-injection dosage and the time-course of Oh (before injection),1.5,3,6and12h of postinjection were as measuring-point for determining the expression level of TLR2, TLR4, IL-1β, TNF-α and IL-6.The results of Real Time-PCR showed that:the expression level of TLR4mRNA in cervix, uterine body and horn in LPS-injection group was lower control group at1.5h,3h and6h after injection, but it was higher than control group at12h after injection. The difference expression level of TLR4between control group and LPS-injection group was not significant at the same time-point after injection (P>0.05), as well as the change of expression level of TLR4in cervix, uterine body and horn with time-course was no significant difference within LPS-injection group (P>0.05). The difference of expression level of TLR2mRNA in cervix, uterine body and horn between control group and LPS group <was not significant at the same point after injection (P>0.05), as well as the change of expression of TLR2in cervix, uterine body and horn with time-course was no significant difference within LPS-injection group (P>0.05).The expression level of IL-1β mRNA in cervix, uterine body and horn in LPS-injection group increased the highest at1.5h after injection and gradually decreased from3to12h after LPS-injection. The difference of expression level of IL-1β mRNA in uterine body and horn between control group and LPS-injection group was significant by1.5h of postinjection (P<0.05),but the change of expression level of IL-1β mRNA in cervix, uterine body and horn was no significant difference with time-course within LPS-injection group (P>0.05). Difference of expression level of IL-6in cervix, uterine body and horn between control group and LPS-injection group was significant by1.5h of postinjection. In cervix, uterine body and horn, the expression level of IL-6mRNA significantly increased at1.5-3h after LPS-injection (P<0.05), and it reached the highest by3h of postinjection. Moreover, the expression level of IL-6at6-12h was significantly lower than1.5-3h in LPS-injection group (P<0.05). The expression level of TNF-a mRNA in cervix, uterine body and hornalso increased the highest1.5h of LPS postinjection and then decreased from3to12h after injection. The difference of expression level of TNF-a mRNA in uterine horn between control group and LPS-injection group was significant at3h after injection (P<0.05). The expression level of TNF-a mRNA significantly increased by1.5or3h of LPS postinjection (P<0.05), moreover it decreased significantly by6-12h (P<0.05).The results suggested that LPS could not promote the expression of TLR2and TLR4in cervix, uterine body and horn of rabbit. While LPS could induce the expression of IL-1β, IL-6and TNF-a in cervix, uterine body and horn of rabbit significantly, and the expression of the genes increased in mammer of time-dependent effect, the expression level of IL-1βand TNF-α increased the highest by1.5h but IL-6did by3h of LPS postinjection. |
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