| Part IThe effect of acupuncture on CCL2, CXCL8and their receptors in rat endometrium of embryo implantation failureObjectiveExplore effect of acupuncture on CCL2, CXCL8and their receptors in rat endometrium with embryo implantation failure.Methods72pregnant rats were obtained through mating between male and female wistar rats. These pregnancy rats were randomly divided into four groups:the normal group (N), the model group (M), acupuncture treatment group (A), and western medicine treatment group (W). Then each group was equally randomized into D6group, D8group, D10group. The rats were treated with mifepristone-sesame oil solution on D1in groups M, group A, and group W, while group N were injected with corresponding sesame oil. Meanwhile,’Housanli’and’Sanyinjiao’were selected for acupuncture. From D1to the time of death, group A were fasten up and then manipulate acupuncture. Corresponding, Group N and group M were only fixed, group W were given progestin. The number of every implanted embryo was calculated. The expression of CCL2, CXCL8and their receptors in rat endometrium were detected by immunohistochemistry, western blot and real-time PCR.Result1. Compared to group N, the average number of implanted embryo were notably decreased in group M (P<0.05); Compared to group M, the average number of implanted embryo was notably increased in group A and W (P<0.05).2. The result of immunohistochemistry:compared to group N, the protein expression of CCL2, CCR2, CXCL8and CXCR1(D6, D8and D10), CXCR2(D8and D10) were notably decreased in rat endometrium of group M (P<0.05). Compared to group M, the protein expression of CCL2and CCR2(D6, D8and D10), CXCL8, CXCR1and CXCR2(D8and D10) were notably increased in rat endometrium of group A(P<0.05); The protein expression of CCL2, CCR2, CXCL8and CXCR1(D6, D8and D10), CXCR2(D8and D10) were remarkably increased in rat endometrium of group W (P<0.05).3. The result of western blot:compared to group N, the protein expression of CCL2, CCR2, CXCL8and CXCR1(D6, D8and D10), CXCR2(D8and D10) were notably decreased in rat endometrium of group M (P<0.05). Compared to group M, the protein expression of CCL2(D6, D8and D10), CCR2and CXCR1(D6and D10), CXCL8and CXCR2(D8and D10) were notably increased in rat endometrium of group A (P<0.05); The protein expression of CCL2, CCR2and CXCR1(D6, D8and D10), CXCL8and CXCR2(D8and DIO) were remarkably increased in rat endometrium of group W (P<0.05).4. The result of real-time PCR:compared to group N, the mRNA expression of CCL2, CCR2and CXCR1(D6, D8and D10), CXCL8and CXCR2(D8and D10) were notably decreased in rat endometrium of group M (P<0.05). Compared to group M, the mRNA expression of CCL2, CXCL8, CXCR1and CXCR2(D8), CCR2(D6and D8) were notably increased in rat endometrium of group A (P<0.05); The mRNA expression of CCL2and CXCL8(D8and D10), CCR2(D6and D8), CXCR1and CXCR2(D8) were remarkably increased in rat endometrium of group W (P<0.05).ConclusionAcupuncture can improve the number of implanted embryos in rats of embryo implantation failure, which may be relevant with up-regulating the expression of CCL2, CCR2, CXCL8, CXCR1and CXCR2at maternal-fetal interface of rats with embryo implantation dysfunction. Part ⅡThe effect of acupuncture on uNK cells, γδT cells and Treg cells in rat of embryo implantation failureObjectiveExplore effect of acupuncture on uNK cells, γδT cells and Treg cells in rat of embryo implantation dysfunction.Methods144pregnant rats were obtained through mating between male and female wistar rats. These pregnancy rats were randomly divided into four groups:the normal group (N), the model group (M), acupuncture treatment group (A), and western medicine treatment group (W). Then each group was equally randomized into D6group, D8group, D10group. The rats were treated with mifepristone-sesame oil solution on D1in groups M, group A, and group W, while group N were injected with corresponding sesame oil. Meanwhile,’Housanli’and’Sanyinjiao’were selected for acupuncture. From D1to the time of death, group A were fasten up and then manipulate acupuncture. Corresponding, Group N and group M were only fixed, group W were given progestin. The proportion ofγδT cells, CD4+CD25+Foxp3+Treg cells in Peripheral blood and CD3-CD161+NK cells, γδT cells, CD4+Foxp3+Treg cells in rat endometrium were detected by flow cytometry. The level of Foxp3protein and mRNA were observed by western blot and real-time PCR, respectively.Result1. Compared to group N, the average number of implanted embryo was remarkably decreased in group M (P<0.05); Compared to group M, the average number of implanted embryo were notably increased in group A and W (P<0.05).2. Compared to group N, the percentage of γδT cells and CD4+CD25+Foxp3+Treg cells were importantly decreased in rat peripheral blood of group M on D6, D8and D10(P<0.05). Compared to group M, the percentage of CD3+γδTCR+cells on D6and D8, CD4+CD25+Foxp3+Treg cells on D6, D8and D10were importantly increased in rat peripheral blood of group A (P<0.05); the percentage of CD3+γδTCR+cells and CD4+CD25+Foxp3+Treg cells were importantly increased in rat peripheral blood of group W on D6, D8and D10(P<0.05).3. Compared to group N, the percentage of CD3"CD161+NK cells, CD3+yγδTCR+cells and CD4+Foxp3+Treg cells were importantly decreased in rat endometrium of group M on D6, D8and D10(P<0.05). Compared to group M, the percentage of CD3-CD161+NK cells and CD3+γδTCR+cells on D6and D8, CD4+Foxp3+Treg cells on D8and D10were importantly increased in rat endometrium of group A (P<0.05); the percentage of CD3-CD161+NK cells and CD4+Foxp3+Treg cells on D6, D8and D10, CD3+y8TCR+cells on D6and D8were importantly increased in rat endometrium of group W (P<0.05).4. Compared to group N, protein expression of Foxp3were remarkably decreased in rat endometrium of group M on D6, D8and D10(P<0.05). Compared to group M, protein expression of Foxp3was remarkably increased in rat endometrium of group A on D6and D8(P<0.05); protein expression of Foxp3was remarkably increased in rat endometrium of group W on D6, D8and D10(P<0.05).5. Compared to group N, mRNA expression of Foxp3were remarkably decreased in rat endometrium of group M on D6, D8and D10(P<0.05). Compared to group M, mRNA expression of Foxp3was remarkably increased in rat endometrium of group A on D6and D10(P<0.05); mRNA expression of Foxp3was remarkably increased in rat endometrium of group W on D6and D8(P<0.05).ConclusionAcupuncture can up-regulate the ratio of y8T cells and CD4+CD25+Foxp3+Treg cells in rat peripheral blood, but also improve the percentage of CD3-CD161+NK cells, γδT cells and CD4+Foxp3+Treg cells in rat endometrium of embryo implantation failure, which indicates that acupuncture can promote embryo implantation and placentation by up-regulation of immune cells. |
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