| BackgroundEndometrium as a highly regenerative tissue in the female can occur cyclicalchanges under the action of estrogen and progesterone. Estrogen can stimulate thegrowth of the endometrium, under the control of the hypothalamus-pituitary-ovarianaxis. When some reasons lead to endometrial lesions, result in uterine front and backwall adhesion and uterine cavity atresia, can be caused by intrauterineadhesions(IUA). In recent years, with the increase of the operation of artificialabortion, drug abortion and intrauterine operation, IUA has become one of thecommon diseases of gynecology and the important causes of infertility, miscarriageand premature delivery. The cure rate of intrauterine is low, after operativesegregation, the adhesion is easy to relapse, which affect the majority of women，sbody and mental health. Therefore, after transcervical resection of adhesion, thetreatment of how to prevent adhesion relapse, promote endometrial growth is a bigproblem in clinical work. At present, after transcervical resection of adhesion,artificial cycle treatment for2~3months is popularly adopted to improve theregeneration of endometrium and prevent adhesion relapse. At present, the dosage ofestrogen and treatment course are not clear, they are used mainly based on patient，sadhesion degree, type and the operator，s experience, and the effect is not satisfactory, the growth of endometrium could not be improved well. Some scholars think that ifthe estrogen physiological dosage is invalid to endometrium, more amount ofestrogen dosage can be added to promote the growth of endometrium. There are otherstudies finding that if the endometrium basal layer is damaged seriously, theregeneration capacity of endometrium reduces, high estrogen level may cause the riseof some fibrosis cytokines, and on the contrary, it can promote the formation of IUA.The effect of different dosage of estrogen and the application time on the growth ofendometrium is not clear now.Infection on the growth of the endometrium is currently not clear, some scholarsbelieve that there is no need to use antibiotics to prevent the operation afterintrauterine operation. But once the infection, the infected inflammatory reaction cancause the aggravation of endometrium damage, leading to endometrium repairdisorder, accelerating the reformulation of intrauterine adhesion. Among allpro-inflammatory cytokines, prostaglandinE2(PGE2) is an important cell growth andregulation factor, has a regulating function to the proliferation and biological functionof multiple cells, it is closely related to the physiology and pathology of endometrium.It can induce the whole body inflammatory reaction, enhance the synthesis ofcollagen biont in cell, and promote the fibrosis change of inflammatory area. But theeffect of prostaglandin E2on endometrial growth and how to adjust to the cells ofendometrium role is unclear.ObjectiveThrough endometrium stromal cell culture in vitro passage system, the topicapplied different dosages and different time estradiol and inflammatory mediatorprostaglandinE2to act on the cell, MTT method and Real-time PCR method wereused to observe the effect of estradiol and inflammatory mediator PGE2on the growthof endometrium stromal cell, the paper provided one early experimental basis forfurther explore the assisted estrogen after transcervical resection of adhesion and ifthe inflammation has an effect on endometrial growth. Material and Methods1. Collagenase digestion method and screening method were used to isolatedculture human endometrium stromal cell, MTT method was used to draw the normalgrowth curve of cell, observe cell growth cycle. Morphology andimmunohistochemistry method were used to identify the cultured cell.2. Human endometrium stromal cell was divided into control group: normalculture solution was added. Estradiol group: estradiol with concentration of1x10-9,10-8,10-7,10-6mol/L were added respectively. Estradiol+PGE2group: estradiol withdifferent concentration(10-9,10-8,10-7,10-6mol/L) and PGE2(50ng/L concentration)were added respectively at the same time. PGE2group was just added by PGE2(50ng/L concentration). MTT method was used to indirectly measure the situation ofendometrium stromal cell proliferation of each group. Real-time PCR method wasapplied to measure the change of intracellular PCNA mRNA expression in eachgroup.3. All experimental data were showed by mean±standard deviation (x±S).Independent sample，s analyze was used by t-test, one-way ANOVA was used toanalyze the data in each group, LSD-t test method was used to pairwisecomparison.Different time points in each group were compared using repeatedmesurement design, test level α=0.05.Results1. Cell morphology features: the stromal cells produced from culture mostly werepolygon, the two ends of cell had many short protuberances, cell arrangement wasnon-polar, and cell grew with adherence. With cellular subculture going, cellgradually stretched into fusiform, had fibroblast morphology.2. Cell growth cycle and immunohistochemistry result:cell growth cycle wasgenerally7-10day, the first day was latent growing phase, the2-4th day entered intoexponential phase, after5-6days, it reached plateau phase, after the7th day, itgradually atrophied and faded away. The specific vimentin antibody dye of stromalcell was positive, positive cell cytoplasm showed yellow color, keratin antibody dye was negative.3. With the action time going, the reproduction rate of stromal cell in estradiolincreased obviously, cell proliferation showed time dependence, the difference hadstatistical significance (P<0.05), there was no difference in the concentration of eachgroup (P>0.05). Compared24h estradiol+PGE2group with E2group, the averagereproduction rate of cell increased, the difference had statistical significance (P<0.05).Compared with72h estradiol+PGE2group, although the cell had growth trend withthe action time going, the cell growth rate decreased obviously when compared withestradiol group (P<0.05). As the extension of the time, cell proliferation rate declinedobviously with the PGE2group(P<0.05).4. With the action time going, the relative expression amount of cell PCNA mRNA in72h estradiol group increased obviously, compared with48h,24h, the difference hadstatistical significance (P<0.05), but there was no obvious difference in theconcentration of each group (P>0.05).24h estradiol group+PGE2group wascompared with estradiol group, the relative expression amount of PCNA mRNAincreased obviously (P<0.05). The relative expression amount of cell PCNA mRNAof72h estradiol group+PGE2group had growth with the action time going, butcompared with estradiol group, the relative expression amount of cell PCNA mRNAdecreased obviously (P<0.05). As the extension of the time, the relative expressionamount of cell PCNA mRNA declined obviously with the PGE2group(P<0.05).Conclusion1. Human endometrium stromal cell was successfully isolated culture in vitro,the morphology of cell and vimentin dye positive met the features of endometriumstromal cell.2. Estradiol can promote the proliferation of endometrium stromal cell, butproliferative effect was not concentration dependency, it was time dependency.3. Inflammatory mediator PGE2can regulate the growth of endometrial cells. |
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