Objective To investigate the effects of repeated high frequency transcranial magnetic stimulation (rTMS) on spatial learning and memory function,and on long-term potentiation (LTP) after global cerebral ischemia and reperfusion,and to explore the mechanisms involved.Methods Eighty-three male Wistar rats were studied.Five were tested to determine their average motor threshold (Tm).The others were divided into a normal control group,a cerebral ischemia and reperfusion model group and an rTMS group.Cerebral ischemia was induced with the four vessel occlusion method for 10 minutes.The rTMS treatment protocol (10 Hz stimulation for 5 s at the resting threshold,twice a day) was applied over a 2-week period from day 3 post-operation.The Morris water maze test was performed to observe spatial learning and memory at post-operation day 2 and day 4.The field excitatory postsynaptic potentials,population spike and the magnitude of long-term potentiation (LTP) induced by theta burst electric stimulation were recorded from the perforant path to the dentate gyrus (PP-DG).Results At post-operation day 3,rats in the untreated cerebral ischemia and reperfusion model group exhibited a significant decrease in the magnitude of the PP-DG LTP as compared to the normal group.No significant difference in LTP was found between the model group and the rTMS group.After the 2 weeks of treatment the LTP levels in the rTMS treated group were significantly higher than in the two untreated groups.In the Morris water maze testing,the average escape latency in the rTMS group was significantly shorter than that of the cerebral ischemia and reperfusion model group (which was not treated).In the probe trials,the time in the original quadrant of the platform and the time of crossing the platform were both significantly less for the rTMS-treated rats than for those not treated.Conclusions High frequency rTMS can improve spatial learning and memory after global cerebral ischemia and reperfusion by enhancing the LTP induced in the hippocampus.High frequency rTMS might exert this beneficial effect by modulating the function of intermediate neurons in the hippocampal neuronal network and by promoting neurotransmitter release.

Objective To investigate the effects of functional electrical stimulation (FES) on motor function and on the expression, proliferation, migration and differentiation of endogenous neural stem cells in the subventricular zone (SVZ) after cerebral ischemia.Methods Middle cerebral artery occlusion (MCAO) was used to induce a model of cerebral ischemia in 108 rats using the modified Zea-Longa method of intraluminal filament occlusion.They were then randomly divided into an FES group, a placebo stimulation group and a control group with 36 cases in each.Superficial FES electrodes were pasted on the paralyzed forelimbs of the rats in the first two groups, though FES treatment was administered only to the FES group beginning on the 3rd day after the MCAO operation.The stimulation was designed to produce extension of the wrist and digits of the paralyzed forelimb.Before, and after 1,3, 7 and 14 days of the treatment, the neurological deficit was evaluated using modified neurological severity scoring (mNSS).BrdU +/GFAP+, BrdU+/DCX+ and BrdU+/NeuN + cells in the SVZ were detected using immunofluorescence technique.Results After 7 and 14 days of treatment, the average motor function of the rats in the FES group had improved significantly when compared with the averages of the other two groups.Compared with the other two groups, the average number of BrdU +/GFAP+ positive cells in the ischemic SVZ was also significantly greater in the FES group after 7 and 14 days of treatment.After 14 days, BrdU +/Dcx + positive cells in the FES group had also increased significantly more,but only a few BrdU +/NeuN + cells had appeared in any of the three groups.Conclusion FES can improve motor function after cerebral ischemia, and promote proliferation and differentiation of neural stem cells in the SVZ.

Objective Interleukin-21(IL-21) is likely to contribute to the development of liver fibrosis, but up to now, no study has been reported on the relationship between IL-21 and intrauterine adhesions ( IUA) .This study aimed to establish a rat model of IUA induced by mechanical injury and lipopolysaccharide (LPS) infection (dual injury), determine the expression level of serum IL-21, and confirm the association of serum IL-21 with the formation of IUA. Methods Forty healthy female SD rats were randomly divided into four groups of equal number,control, mechanical injury, LPSinfection, and dual injury.At 7 days after IUA modeling,uterine tissue-swere collected from all the animals for observation of the endometrial glands,detection of the degree of IUA by Masson staining, measure-ment of the serum IL-21 level by radioimmunoassay, and analysis of the correlation between the number of endometrial glands and the de-gree of fibrosis. Results The number of endometrial glandswas significantly smallerin the dual injury group (3.59±1.20) than in the mechanical injury (11.66±2.34) and LPSinfection group(11.59±1.47)(P<0.05), while the proportion of fibrosis area wassignificantly higher in the former group(0.65±0.03) than in the lattertwo(0.30±0.07 and 0.32±0.08)(P<0.05).The level of serum IL-21 was signifi-cantly increased in the dual injury group ([286.21±27.80]pg/mL) as compared with those in the control ( [ 118.65 ±22.55 ] pg/mL ) , mechanical injury([176.20±19.05]pg/mL), and LPS infection group ([187.98±16.51]pg/mL) (P<0.05), with a positive correlation be-tween the IL-21 level and theproportion of fibrosis area ( r=0.271, P<0.05) . Conclusion A rat model ofIUAwas successfully established by mechanical injury and lipopolysaccharide (LPS) infection.The evi-dent increase of serum IL-21 in the IUA model was positively correlated with the percentage of fibrosis area, suggesting that IL-21 may be involved indirectly in the formation of IUA.

Objective To investigate the effective administration model for gestational diabetes mellitus (GDM) in the community. Methods In a prospective study, the 75 g oral glucose tolerance test (OGTT) was performed in 4 713 resident pregnant women over 20 years old who received antenatal care in a general hospital or a special hospital from Sep. 2011 to Aug. 2012. Five hundred and thirty-three pregnant women were diagnosed as GDM, 198 patients who labored in a general hospital were enrolled in thegroup A , and the rest who labored in a special hospital were enrolled in the group B. 198 cases with non-GDM were enrolled in the group C. Results The incidence of GDM during this study period was 11.3%. The maternal age , gestationl weeks and OGTT results of patients in the three groups were significantly different (P < 0.05). With the increase of maternal age of pregnant women, the blood glucose increased while the gestationl weeks descreased. No significant differencees were shown in prevalence of polyhydramnios, hypertensive disorder complicating pregnancy, premature labor, macrosomia , fetal growth restriction and neonatal asphyxia among the three groups (P > 0.05). However, significant differences were found in the incidence of postpartum hemorrhage and cesarean section. The incidence of postpartum hemorrhage increased significantly in the patients of group B (χ2= 7.156, v = 2, P = 0.028). The incidence of cesarean section increased significantly in the patients of group A (χ2= 63.592, v = 2, P = 0.000). Conclusion Establishing an effective administration model for gestational diabetes mellitus in the community could control the incidence of GDM associated complications.

AIM:To investigate the differentiation from rat mesenchymal stem cells (rMSC) into neuron-like cells. METHODS:rMSC were separated from femur marrow and expanded in L-DMEM culture medium supplemented with 10% FSC. rMSC were induced to differentiate into neurons with L-DMEM/adrenaline,L-DMEM/noradrenaline and L-DMEM/isoprenaline, respectively. Meanwhile, rMSC were cultured in L-DMEM in control group. Nestin, neuron-specific enclose (NSE), glial fibrillary acidic protein (GFAP) were detected by immunocytochemistry. RESULTS: rMSC were expanded as undifferentiated cells in culture from 5 to 22 passages, indicating their differentiated capacity. Simple method induced rMSC to exhibit a neuronal phenotype, expressing positive NSE,nestin, and GFAP, at 5 hours in all group. The undifferentiating cells (control group 53.1%?4.3%), and differentiating cells (treated group: adrenaline 74.7%?2.6%; noradrenaline 75.9%?2.4%; isoprenaline 72.1%?4.4%), expressed characteristics of various neuronal cells, from 5 hours to 6 days. There were neuron-like cells in rMSC cultured in L-DMEM/10%FBS from 7 to 13 passage(66.5%?6.4%). CONCLUSION: It suggests that rat neural stem cells (rNSC) exist in bone marrow, rMSC can be differentiated into various neural cells with adrenaline hormones in vitro.

Objective To investigate the effects of functional electrical stimulation (FES) on motor function and the expression of bromodeoxyuridine (Brdu) + and glial fibrillary acid protein (GFAP) + in the subventricular zone (SVZ) of rats with acute cerebral infarction,and to explore it's mechanism. Methods A rat model of cerebral infarction was established using Longa's technique for middle cerebral artery occlusion (MCAO) with an intraluminal filament.The rats were randomly divided into a FES group,a placebo stimulation group and a control group.In each group,rats were randomly allocated into 1 d,3 d,7 d and 14 d subgroups (6 rats/subgroup).Superficial electrodes were pasted on the paralyzed forelimbs of rats in the FES group for connecting with the FES instrument,and FES treatment was carried out with a current of 4-5 mA for 15 min on the third day after the MCAO operation to produce extension of the wrist and the digits of the paralyzed forelimb.The rats in the placebo stimulation group were pasted with electrodes,but no FES was administered and they received no other treatment.Neurological deficits were evaluated using the modified neurological severity score (mNSS) before treatment and on the 1 st,3rd,7th,and 14th day after treatment. BrdU and GFAP positive cells in the SVZ were detected by immunofluorescence techniques.Results After 7 or 14 days the motor function of rats in the FES group had improved significantly compared with the placebo stimulation and control groups.Compared with the other two groups,the expression levels of BrdU+ and GFAP+ cells in the ischemic SVZ in the FES group were significantly higher at the 3rd,7th and 14th day.Conclusion FES can improve motor function after acute cerebral infarction and also promote the proliferation and differentiation of neural stem cells in the SVZ.

BACKGROUND: By targeting inducing differentiation in vitro,mesenchymal.stem cells(MSCs) transform into osteoblasts,lipocytes,chondrocytes,muscular cells,neuronal cells,etc. Whether Chinese herbs act on induced differentiation of MSCs in rats or not?OBJECTIVE: To study the amplification of MSCs cultured in vitro in SD rats and efficacy of Chinese herbs on targeting inducing differentiation of neuron-like cells.DESIGN: Exploring study with repeated observation and measurement based on cells.SETTING: Department of pathophysiology in a medical college.MATERIALS: Experimental marrow collected from SD male tested-healthy rats.METHODS: By adhesion method,MSCs in rats were isolated for amplifying culture in vitro. Flow-type cell instrument was applied for the determination of its surface antigen expression. Various Chinese herbal components were used for the targeting inducing differentiation of MSCs into neuron-like cells. The cellular morphology was observed under optical microscope. and the specific antigen label of neuronal cells was determined with immunocyto-chemical method.MAIN OUTCOME MEASURES:①Results of MSCs isolation and amplification;②Results of identification of MSCs surface antigen and neuon-like cells.RESULTS: By adhesion method,MSCs in rats were isolated successfully and amplified in a large amount in vitro. It was indicated in the results determined by flow-type cell instrument that CD14,CD1 1α,CD34,CD38,CD45,CD80 and CD86 presented negative,and CD29,CD44,CD90,CD105 and CD166 presented positive. By induced with various kinds of Chinese herbs,like huangqi(Radix Astragali seu Hedysari),tianma(Rhizoma Gastrodiae),renshen (Radix Ginseng),danggui(Radix Angelicae Sinensis),naoxinshu,renshen fengwangjian for 1 to 3 hours,most MSCs transformed into neuron-like cells,presenting soma and neurite. With immunocyto-chemical staining,neuron-specific enolase(NSE) and nestin displayed positive and glial fibrillary acidic protein negative.CONCLUSION: MSCs in SD rats have the potential of multi-targeting differentiation,presenting a strong capacity of amplification and self-replacement. In a suitable inducing condition,MSCs may differentiate into neuron-like cells.

Objective To explore the effects of different intrauterine adhesion (IUA) classification systems on predicting the IUA prognosis.Methods One hundred cases were selected as the subjects in present study from those diagnosed with IUA and underwent surgery in Zhujiang Hospital of Southern Medical University from Jan.2010 to Jan.2017,and were followed up for two years.According to the actual situation,all patients were scored by March,AFS,ESGE and Chinese classification for comparing the effects of different IUA classification systems on predicting the pregnancy rate,live birth rate and effective rate within 2 years after surgery.Results ESGE classification had a good effect on predicting the postoperative live birth rate and effective rate,and a certain predictive effect on pregnant rate,with the area under curve (AUC) of 0.722,0.754 and 0.635,respectively.March classification had a certain effect on predicting the postoperative live birth rate and effective rate with AUC of 0.635,0.754,respectively,but had a poor effect on predicting pregnant rate.AFS classification and China classification had poor effect on predicting the IUA prognosis.Conclusion ESGE classification system is better than the other systems including March,AFS and Chinese classification,on predicting the IUA prognosis,but further verification in large sample size is still required.

Objective To investigate the effect of estrogen on the fibrosis process of intrauterine adhesions and the expression of forkhead box F2 (FoxF2).Methods Primary human endometrial stromal cells (HESCs) were obtained by separation with 0.2％ collagenase Ⅰ digestion-mesh filtration-differential adherence,and identified by immunocytochemistry.HESCs affected with 10ng/ml transforming growth factor β1 (TGF-β1) for 48 hours.HESCs in model group were affected with 0,10-6,10-8,10-10 and 10-12mol/L estrogen,the expressions of smooth muscle actin alpha (α-SMA),Collagen I (COL Ⅰ) and FoxF2 were detected by quantitative PCR (qPCR) and Western blotting.Results HESCs with high purity and good activity were obtained by using 0.2％ collagenase Ⅰ digestion-mesh filtration-differential adherence separation method.Immunocytochemistry showed positive vimentin and negative cytokeratin 18 in HESCs.The results of qPCR and Western blotting showed that the mRNA and protein expression levels of α-SMA,COL Ⅰ and FoxF2 were higher in model group than in control group (P＜0.05),the model was built successfully.qPCR revealed that the mRNA expression levels ofα-SMA,COL Ⅰ and FoxF2 were significantly lower in 10-6,10-8 and 10-10mol/L estrogen groups than in model group (P＞0.05 in 10-10mol/L estrogen group,P＜0.05 in other groups),while in 10-12mol/L estradiol group,the expression levels of FoxF2 mRNA significantly decreased (P＜0.05),and of α-SMA and COL Ⅰ mRNA increased,but no significant difference were found (P＞0.05).Compared with the model group,the protein expression levels of α-SMA,COL Ⅰ and FoxF2 in 10-6,10-8 and 10 10mol/L estrogen groups decreased,but no significant difference was found (P＜0.05),while in 10-12mol/L estradiol group,the expression levels ofα-SMA protein increased (P＞0.05),and of COL Ⅰ and FoxF2 proteins decreased (P＜0.05).Conclusions The expression of FoxF2 in intrauterine adhesions is increased.Estrogen can reverse the fibrosis process of intrauterine adhesions in a certain range and inhibit the expression of FoxF2.

OBJECTIVETo investigate the histomorphology and the expressions of the proliferation marker Ki-67 and estrogen receptor in the uterus of mice with autoimmune premature ovarian failure (POF) induced by zona pellucida 3 peptide (pZP3).

METHODSAutoimmune POP models were established in 20 female BALB/c mice (7-8 weeks old) by immunization with pZP3 and another 20 mice served as the control group. The POP models were verified by vaginal cytology, serum sex hormones, ovary histomorphology and ZP3 antibody immunohistochemistry. The histomorphology and expressions of Ki-67, estrogen receptor α and estrogen receptor β in the uterus of the mice were detected.

RESULTSAutoimmune POP models were established successfully in 80% of the mice at 8 weeks after the immunization. Compared with those in the control group, the mice in the model group showed a smaller volume of the uterus, thinner endometrium and a reduced number of glands. The luminal epithelial cells, glandular epithelial cells and stromal cells in the uterus of the model mice all presented with a lower expression of Ki-67 than those in the control group, and Ki-67 translocation from the nuclei to the cytoplasm was found in the model group. The luminal epithelial cells, glandular epithelial cells and stromal cells showed positive ERα immunoreactivity in the model group but not in the control group. No obvious ERβ expression was found in the uterus in either of the groups.

CONCLUSIONpZP3 can induce autoimmune POP, cause suppressed proliferation of the endometrial epithelial cells and stromal cells, and reduce the cellular expression of ERα in the uterus of mice.

Animals ; Autoimmune Diseases ; metabolism ; Cell Nucleus ; Egg Proteins ; Endometrium ; Epithelial Cells ; Estrogen Receptor alpha ; metabolism ; Estrogen Receptor beta ; metabolism ; Female ; Immunohistochemistry ; Ki-67 Antigen ; metabolism ; Membrane Glycoproteins ; Mice ; Mice, Inbred BALB C ; Primary Ovarian Insufficiency ; metabolism ; Receptors, Cell Surface ; Stromal Cells ; Uterus ; metabolism ; Zona Pellucida Glycoproteins