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Objective To investigate the effect of pituitrin-water separation on hemoglobin and ovarian func-tion in laparoscopic ovarian chocolate cyst removal.Methods From February 2018 to February 2019,82 patients with ovarian chocolate cyst removed by laparoscopy were selected from the Second Affiliated Hospital of Wenzhou Medical University.They were randomly divided into observation group and control group according to the digital table,with 41 cases in each group.The control group underwent traditional laparoscopic tear-and-tear ovarian cyst removal,while the observation group underwent pituitrin-water separation.The changes of perioperative indicators ,the decrease of hemoglobin (Hb) before and after operation ,the positive rate of normal ovarian tissue on cyst wall and the number of normal follicles attached to cyst wall ,the changes of serum hormone levels before and after operation were compared between the two groups.Results The amount of bleeding during operation in the observation group [(58.97 ±8.74)mL] was less than that in the control group [(118.93 ±24.21)mL],and the operation time in the observation group[(57.46 ±8.27) min] was shorter than that in the control group [(87.38 ±10.19) min] ,the differences were ststistically significant between the two group ( t ＝14.916,14.598,all P ＜0.05 ).There was no statistically significant difference in the time of anal exhaust between the two groups (P＞0.05).The decrease of Hb in the observation group [(0.71 ±0.16)g／L] was lower than that in the control group [(1.27 ±0.35) g／L] ( t＝9.318,P＜0.05).The positive rate of normal ovarian tissue on cyst wall in the observation group (21.95%) was lower than that in the control group (56.10%),the number of normal follicles attached to the cyst wall in the obser-vation group (2.65 ±0.49) was less than that in the control group (4.86 ±1.24) ,the differences were ststistically significant between the two group ( χ2 ＝10.045, t ＝10.613, all P ＜0.05).The serum level of E2 [( 398.21 ± 17.84)pmol／L] in the observation group was higher than that in the control group [(367.83 ±15.21) pmol／L], while FSH [(6.72 ±0.28)mIU／mL] and LH [(5.23 ±0.38)mIU／mL] levels in the observation group were lower than those in the control group [(7.19 ±0.35)mIU／mL and (5.69 ±0.31)mIU／mL],the differences were ststisti-cally significant between the two group (t＝8.298,6.714,6.006,all P＜0.05).Conclusion The method of pitui-trin-water separation is effective in laparoscopic ovarian chocolate cyst removal ,which can reduce the injury during operation,has little effect on hemoglobin and improve the ovarian reserve function of patients .
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Objective@#To investigate the effect of pituitrin-water separation on hemoglobin and ovarian function in laparoscopic ovarian chocolate cyst removal.@*Methods@#From February 2018 to February 2019, 82 patients with ovarian chocolate cyst removed by laparoscopy were selected from the Second Affiliated Hospital of Wenzhou Medical University.They were randomly divided into observation group and control group according to the digital table, with 41 cases in each group.The control group underwent traditional laparoscopic tear-and-tear ovarian cyst removal, while the observation group underwent pituitrin-water separation.The changes of perioperative indicators, the decrease of hemoglobin (Hb) before and after operation, the positive rate of normal ovarian tissue on cyst wall and the number of normal follicles attached to cyst wall, the changes of serum hormone levels before and after operation were compared between the two groups.@*Results@#The amount of bleeding during operation in the observation group [(58.97±8.74)mL] was less than that in the control group [(118.93±24.21)mL], and the operation time in the observation group[(57.46±8.27)min] was shorter than that in the control group [(87.38±10.19)min] , the differences were ststistically significant between the two group(t=14.916, 14.598, all P<0.05). There was no statistically significant difference in the time of anal exhaust between the two groups (P>0.05). The decrease of Hb in the observation group [(0.71±0.16)g/L] was lower than that in the control group [(1.27±0.35)g/L] (t=9.318, P<0.05). The positive rate of normal ovarian tissue on cyst wall in the observation group (21.95%) was lower than that in the control group (56.10%), the number of normal follicles attached to the cyst wall in the observation group (2.65±0.49) was less than that in the control group (4.86±1.24) , the differences were ststistically significant between the two group(χ2=10.045, t=10.613, all P<0.05). The serum level of E2 [(398.21±17.84)pmol/L] in the observation group was higher than that in the control group [(367.83±15.21)pmol/L], while FSH [(6.72±0.28)mIU/mL] and LH [(5.23±0.38)mIU/mL] levels in the observation group were lower than those in the control group [(7.19±0.35)mIU/mL and (5.69±0.31)mIU/mL], the differences were ststistically significant between the two group (t=8.298, 6.714, 6.006, all P<0.05).@*Conclusion@#The method of pituitrin-water separation is effective in laparoscopic ovarian chocolate cyst removal, which can reduce the injury during operation, has little effect on hemoglobin and improve the ovarian reserve function of patients.
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Objective To explore the effects of Hippo pathway on differentiation, proliferation, and migration of bone marrow mesenchymal stem cells (BMSCs) in vitro. Methods BMSCs of C57BL/6 mice were identified using fluorescence-activated cellsorting analysis and the capabilities of osteogenic, chondrogenic and adipogenic differentiation were evaluated. The differentiation of BMSCs to typeⅡalveolar epithelial cells (AECⅡ) was induced by indirect co-culture with mouse lung epithelial cells (MLE-12) and small airway epithelial cell growth medium (SAGM). The Hippo pathway was regulated by 2-deoxy-D-glucose (2-DG) and 9E1, the effects of 2-DG and 9E1 on the expression of BMSCs surface proteins (SPB, SPC and SPD) mRNA and pro-SPC protein were detected by real time quantitative polymerase chain reaction (qRT-PCR) and Western Blot. The effect of Hippo pathway on differentiation of BMSCs to AECⅡ cells was evaluated. The effect of Hippo pathway on the proliferation of BMSCs was evaluated by methyl thiazolyl tetrazolium (MTT) assay (intervention of 0.1, 0.5, 1.0, 5.0 mmol/L 2-DG). The scratch test and Transwell chamber test were used to analyze the effect of Hippo pathway on migration ability of BMSCs to conditioned medium of acute respiratory distress syndrome (ARDS) lung tissue. Results 2-DG could activate Hippo pathway in a dose-dependent manner and promote the differentiation to AECⅡ and proliferation of BMSCs, the maximum effects were observed after 5 mmol/L of 2-DG treatment [SPB mRNA (2-ΔΔCT): 2.42±0.28 vs. 1.89±0.11, SPC mRNA (2-ΔΔCT): 8.06±0.68 vs. 6.59±0.79, SPD mRNA (2-ΔΔCT): 6.45±0.37 vs. 5.27±0.28, pro-SPC/β-actin: 5.80±1.86 vs. 4.93±1.18, proliferation rate:(145.46±18.18)% vs. (98.91±4.36)%, all P < 0.05], but 9E1 could reverse those effects through inhibition of Hippo pathway [SPB mRNA (2-ΔΔCT): 1.32±0.17 vs. 1.89±0.11, SPC mRNA (2-ΔΔCT): 3.91±0.34 vs. 6.59±0.79, SPD mRNA (2-ΔΔCT): 3.38±0.25 vs. 5.27±0.28, pro-SPC/β-actin: 2.48±0.17 vs. 4.93±1.18, proliferation rate: (80.00±7.27)% vs. (98.91±4.36)%, all P < 0.05]. The ability of horizontal migration [wound healing: (27.17±3.53)% vs. (52.45±6.52)%, P < 0.05] and homing BMSCs to conditioned medium of ARDS lung tissue [cell count (fold, relative to control): 2.77±0.21 vs. 1.90±0.19, P < 0.05] were increased after activation of Hippo pathway by 2-DG treatment, but those effects were reversed after inhibition of Hippo pathway by 9E1 treatment [wound healing: (79.89±8.42)% vs. (52.45±6.52)%, cell count (fold, relative to control): 1.69±0.13 vs. 1.90±0.19, both P < 0.05]. Conclusion Activation of Hippo pathway could enhance differentiation of BMSCs to AECⅡ, promote proliferation and ability of horizontal migration and homing BMSCs to conditioned medium of ARDS lung tissue in vitro.
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Objective To investigate the effects of Hippo signaling pathway on lung injury repair of mesenchymal stem cells (MSC) in acute respiratory distress syndrome (ARDS) and its mechanism. Methods Mouse bone marrow-derived MSC (mMSCs) cell lines with low expression of large tumor suppressor 2 (LATS2) were constructed by lentiviral vector transfection. Male C57BL/6 mice aging 6-8 weeks old were divided into four groups according to random number table (n = 36). The ARDS animal model (ARDS group) was reproduced by intratracheally injection of 2 g/L lipopolysaccharide (LPS) 50 μL, the normal saline (NS) control group was injected with an equal volume of NS. After 4 hours of model reproduction, 5×104 mMSCs transfected with blank lentivirus vector (MSC-shcontrol group) or shLATS2 lentivirus vector (MSC-shLATS2 group) were transplanted intratracheally, while NS control group and ARDS group were injected with equal volume of phosphate buffered saline (PBS). Mice were sacrificed at 3, 7, and 14 days after modeling, and lung tissue and bronchoalveolar lavage fluid (BALF) were harvested. Near-infrared fluorescence imaging, immunofluorescence staining and Western Blot were used to track mMSCs in lung tissue. Retension and proportion of mMSC differentiation into type Ⅱ alveolar epithelial cells (AECⅡ) were evaluated. Lung tissue wet weight/body weight ratio (LWW/BW) and total protein (TP) and albumin (ALB) in BALF were determined to reflect pulmonary edema. The expression of Occludin protein in lung epithelium was tested by Western Blot to reflect permeability of epithelium. The levels of interleukins (IL-1β, IL-6, IL-10) in BALF were assessed by enzyme-linked immunosorbent assay (ELISA) to reflect lung inflammation. Hematoxylin-eosin (HE) staining and modified Masson staining were carried out, and the scores were assessed to reflect lung injury and evaluate pulmonary fibrosis. Results The signal intensity of isolated lung fluorescence images at 3 days in the MSC-shLATS2 group was significantly higher than that in the MSC-shcontrol group (fluorescence intensity: 0.039±0.005 vs. 0.017±0.002, P < 0.05), the number of green fluorescent protein (GFP)-positive cells in lung tissue was also significantly higher than that in the MSC-shcontrol group (cells/HP:29.65±6.98 vs. 17.50±4.58, P < 0.05), but they all decreased with time; and the proportion of mMSCs differentiated into AECⅡ was significantly increased [(64.12±15.29)% vs. (19.64±3.71)%, P < 0.05]. Compared with the NS control group, the levels of surface active protein C (SPC) and Occludin protein in the ARDS group were significantly decreased, LWW/BW ratio and TP, ALB and inflammatory factors levels in BALF were significantly increased; extensive alveolar and interstitial edema, hemorrhage and diffuse inflammatory cell infiltration were found in lung tissue, and the lung injury score was significantly increased; collagen fibers were deposited in alveolar septum and alveolar cavity, and pulmonary fibrosis score was also increased significantly. Compared with the ARDS group, the expression levels of SPC and Occludin at 14 days in the MSC-shcontrol group and the MSC-shLATS2 group were significantly increased (SPC/β-actin: 0.51±0.12, 0.68±0.10 vs. 0.27±0.08, Occludin/β-actin: 0.49±0.19, 0.79±0.11 vs. 0.25±0.08, all P < 0.05), TP, ALB, IL-1β, IL-6 levels in BALF at 3 days were significantly decreased [TP (g/L): 8.08±1.72, 5.12±0.87 vs. 12.55±2.09; ALB (g/L): 0.71±0.21, 0.44±0.18 vs. 1.18±0.29, IL-1β(ng/L): 99.26±14.32, 60.11±8.58 vs. 161.86±25.17, IL-6 (ng/L): 145.54±13.29, 101.74±11.55 vs. 258.79±27.88, all P < 0.05], and IL-10 was significantly increased (ng/L: 190.83±22.61, 316.65±37.88, both P < 0.05). Furthermore, all the above parameters in the MSC-shLATS2 group were significantly improved as compared with those in the MSC-shcontrol group (all P < 0.05). LWW/BW ratio in the MSC-shLATS2 group was significantly lower than that in the ARDS group and the MSC-shcontrol group (mg/g: 9.85±1.51 vs. 16.78±1.92, 14.88±1.74, both P < 0.05). Conclusion Inhibiting Hippo signaling pathway by low expression of LATS2 could promote the retention of mMSCs in lung tissue and differentiation into AECⅡcells of ARDS mice, improve pulmonary edema and alveolar epithelial permeability, regulate pulmonary inflammatory response, and alleviate pathological damage and fibrosis of lung tissue.
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Objectives To determine the effect of Hippo signaling pathway on lung injury repair of bone-marrow derived mesenchymal stem cells (mMSCs) in murine lipopolysaccharide (LPS) induced acute respiratory distress syndrome (ARDS).Methods C57BL/6 mouse bone marrow-derived MSC (mMSCs) cell lines with low expression of large tumor suppressor 1 (LATS 1) were constructed by lentiviral vector transfection.ARDS was modeled by intratracheally injection of 2 mg/mL lipopolysaccharide (LPS)50 μL.C57BL/6 mice were randomly(random number) divided into four groups (n=36):normal control group,ARDS group,ARDS mice transplanted with mMSCs transfected with blank lentivirus vector (MSC-shcontrol group) or sh-LATS 1 lentivirus vector (MSC-shLATS 1 group).Mice were sacrificed at 3,7,and 14 d after modeling,and lung tissue and bronchoalveolar lavage fluid (BALF) were collected.Near-infrared fluorescence imaging,immunofluorescence staining and Western blot were used to evaluate retention and differentiation of mMSCs in lung tissue.Lung tissue wet weight/body weight ratio (LWW/BW) and total protein (TP) and albumin (ALB) in BALF were determined to reflect pulmonary edema.The expression of Occludin protein in lung epithelium was tested by Western blot.The levels of interleukins (IL-1β,IL-6,and IL-10) in BALF were assessed by enzyme-linked immunosorbent assay (ELISA).Lung injury score and pulmonary fibrosis score in lung tissue were assessed.Results The retention ofmMSCs at 3,7 and 14 d in the MSC-shLATS1 group were significantly higher than those in the MSC-shcontrol group [(26.25±4.58) vs (12.13±3.75) cells/HP,(20.49±3.86) vs (9.97±2.76) cells/HP,and (13.77±3.55) vs (6.89±2.10) cells/HP.all P＜0.05],so was the differentiation of mMSCs into type Ⅱ alveolar epithelial cells at 14 d [(64.12±15.29)％ vs (19.64±3.71)％,P＜0.05].LWW/BW and TP and ALB in BALF at 3 and 14 d in the MSC-shLATS1 group [(9.85±1.51),(6.11±0.83) (mg/g) and (5.12±0.87),(3.05±0.87) (mg/mL)and (0.44±0.18),(0.33±0.04) (mg/mL)] were higher than those in the MSC-shcontrol group [(14.88± 1.74),(8.04±l.70)(mg/g) and (8.08±1.72),(5.94±1.20) (mg/mL) and (0.71±0.21),(1.07±0.29) (mg/mL)] (all P＜0.05),so was the relative expression of Occludin protein[(0.79±0.11) vs (0.49±0).19),(P＜0.05)].The levels of IL-1β and IL-6(pg/mL) in BALF in the MSC-shLATS1 group [(60.11±8.58),(101.74±11.55)] was lower than those in the MSC-shcontrol group [(99.26±14.32),(145.54±13.29)] (all P＜0.05),but the levels of IL-10 in BALF in the MSC-shLATS 1 group (316.65±37.88)pg/mL was higher than those in the MSC-shcontrol group (190.83±22.61)pg/mL (P＜0.05).Lung injury scores at 3 and 14 d in the MSC-shLATS1 group [(7.18±1.12),(3.33±0.49)] was lower than those in the MSC-shcontrol group [(9.72±1.45),(5.11±0.86)] (all P＜0.05),so was pulmonary fibrosis score at 14 d [(0.68±0.12) vs (1.47±0.18),P＜0.05].Conclusion Inhibition of Hippo signaling pathway through underexpression of LATS1 could improve the therapeutic effects of mMSCs in murine LPS-induced ARDS.
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OBJECTIVE@#To investigate the effects of Hippo signaling pathway on lung injury repair of mesenchymal stem cells (MSC) in acute respiratory distress syndrome (ARDS) and its mechanism.@*METHODS@#Mouse bone marrow-derived MSC (mMSCs) cell lines with low expression of large tumor suppressor 2 (LATS2) were constructed by lentiviral vector transfection. Male C57BL/6 mice aging 6-8 weeks old were divided into four groups according to random number table (n = 36). The ARDS animal model (ARDS group) was reproduced by intratracheally injection of 2 g/L lipopolysaccharide (LPS) 50 μL, the normal saline (NS) control group was injected with an equal volume of NS. After 4 hours of model reproduction, 5×104 mMSCs transfected with blank lentivirus vector (MSC-shcontrol group) or shLATS2 lentivirus vector (MSC-shLATS2 group) were transplanted intratracheally, while NS control group and ARDS group were injected with equal volume of phosphate buffered saline (PBS). Mice were sacrificed at 3, 7, and 14 days after modeling, and lung tissue and bronchoalveolar lavage fluid (BALF) were harvested. Near-infrared fluorescence imaging, immunofluorescence staining and Western Blot were used to track mMSCs in lung tissue. Retension and proportion of mMSC differentiation into type II alveolar epithelial cells (AEC II) were evaluated. Lung tissue wet weight/body weight ratio (LWW/BW) and total protein (TP) and albumin (ALB) in BALF were determined to reflect pulmonary edema. The expression of Occludin protein in lung epithelium was tested by Western Blot to reflect permeability of epithelium. The levels of interleukins (IL-1β, IL-6, IL-10) in BALF were assessed by enzyme-linked immunosorbent assay (ELISA) to reflect lung inflammation. Hematoxylin-eosin (HE) staining and modified Masson staining were carried out, and the scores were assessed to reflect lung injury and evaluate pulmonary fibrosis.@*RESULTS@#The signal intensity of isolated lung fluorescence images at 3 days in the MSC-shLATS2 group was significantly higher than that in the MSC-shcontrol group (fluorescence intensity: 0.039±0.005 vs. 0.017±0.002, P < 0.05), the number of green fluorescent protein (GFP)-positive cells in lung tissue was also significantly higher than that in the MSC-shcontrol group (cells/HP: 29.65±6.98 vs. 17.50±4.58, P < 0.05), but they all decreased with time; and the proportion of mMSCs differentiated into AEC II was significantly increased [(64.12±15.29)% vs. (19.64±3.71)%, P < 0.05]. Compared with the NS control group, the levels of surface active protein C (SPC) and Occludin protein in the ARDS group were significantly decreased, LWW/BW ratio and TP, ALB and inflammatory factors levels in BALF were significantly increased; extensive alveolar and interstitial edema, hemorrhage and diffuse inflammatory cell infiltration were found in lung tissue, and the lung injury score was significantly increased; collagen fibers were deposited in alveolar septum and alveolar cavity, and pulmonary fibrosis score was also increased significantly. Compared with the ARDS group, the expression levels of SPC and Occludin at 14 days in the MSC-shcontrol group and the MSC-shLATS2 group were significantly increased (SPC/β-actin: 0.51±0.12, 0.68±0.10 vs. 0.27±0.08, Occludin/β-actin: 0.49±0.19, 0.79±0.11 vs. 0.25±0.08, all P < 0.05), TP, ALB, IL-1β, IL-6 levels in BALF at 3 days were significantly decreased [TP (g/L): 8.08±1.72, 5.12±0.87 vs. 12.55±2.09; ALB (g/L): 0.71±0.21, 0.44±0.18 vs. 1.18±0.29, IL-1β (ng/L): 99.26±14.32, 60.11±8.58 vs. 161.86±25.17, IL-6 (ng/L): 145.54±13.29, 101.74±11.55 vs. 258.79±27.88, all P < 0.05], and IL-10 was significantly increased (ng/L: 190.83±22.61, 316.65±37.88, both P < 0.05). Furthermore, all the above parameters in the MSC-shLATS2 group were significantly improved as compared with those in the MSC-shcontrol group (all P < 0.05). LWW/BW ratio in the MSC-shLATS2 group was significantly lower than that in the ARDS group and the MSC-shcontrol group (mg/g: 9.85±1.51 vs. 16.78±1.92, 14.88±1.74, both P < 0.05).@*CONCLUSIONS@#Inhibiting Hippo signaling pathway by low expression of LATS2 could promote the retention of mMSCs in lung tissue and differentiation into AEC II cells of ARDS mice, improve pulmonary edema and alveolar epithelial permeability, regulate pulmonary inflammatory response, and alleviate pathological damage and fibrosis of lung tissue.
Assuntos
Animais , Masculino , Camundongos , Via de Sinalização Hippo , Lesão Pulmonar/prevenção & controle , Células-Tronco Mesenquimais/metabolismo , Camundongos Endogâmicos C57BL , Proteínas Serina-Treonina Quinases/metabolismo , Síndrome do Desconforto Respiratório do Recém-Nascido/metabolismo , Transdução de SinaisRESUMO
Objective To evaluate the prognostic value of transcutaneous oximetry in patients with septic shock.Methods Fifty-three patients with septic shock were enrolled prospectively from January 2013 to December 2015.Transcutaneous oximetry were used to determine the results of 10 min oxygen challenge tests (OCT) carried out at beginning(0 h) and at 6 h after fluid resuscitation respectively.The 10-min OCT value (10 min OCT) and oxygen challenge index(OCI) were calculated.The APACHE Ⅱ and SOFA score,hemodynamic variables,oxygen metabolism indexes,dose of vasoactive agents,10 min OCT,and OCI at 0 h and at 6 h were recorded.Patients were assigned into survival group and death group according to the 28 d survival.The differences in demographics and clinical data were compared between groups.The role of 10 min OCT and OCI in predicting death was evaluated by receiver operating characteristic curves(ROC).The Kaplan-Meier surviving curve was created and the survival of the patients was analyzed by the Log-rank test.Risk factors associated with the prognosis were analyzed using the multiple logistic regression analysis.Results There were 29 patients in the survival group and 24 patients in the death group.Compared with death group,10 min OCT[(77.55±18.48)mmHg vs.(51.30±21.60)mmHg] and OCI [(0.78±0.13) vs.(0.59±0.15)] at 6 h in survival group were significantly higher(P＜0.05),while APACHE Ⅱ [(12.48±5.69) vs.(17.25±8.79)] and SOFA [(5.79±1.72) vs.(10.10±2.52)] in survival group were significantly lower than those in death group(P＜0.01).The area under the ROC curve of 10 min OCT at 6 h and OCI at 6 h for predicting 28 d death were 0.86±0.05(95％CI:0.76-0.87,P＜0.01) and 0.79±0.08(95％CI:0.64-0.95,P＜0.01),respectively.The optimal cutoff point for 10 min OCT at 6 h was 72.00 mmHg with the sensitivity of 76.84％ and specificity of 85.03％.The optimal cutoff point for OCI at 6 h was 0.76 with the sensitivity of 76.84％ and specificity of 77.47％.Kaplan-Meier survival analysis showed that 28 d survival rate in high level of 10 min OCT at 6 h and high level of OCI at 6 h were significantly higher than that in low level of 10 min OCT at 6 h(70.86％ vs.31.82％,x2=7.96,P＜0.01)and low level of OCI at 6 h (75.00％ vs.32.00％,x2=9.86,P＜0.01).Multivariate logistic regression analysis showed that both 10 min OCT at 6 h (OR=0.92,95％CI:0.88-0.96,P＜0.05) and OCI at 6 h (OR=0.01,95％CI:0.001-0.023,P＜0.05) were independent risk factors associated with 28 d mortality of patients with septic shock.Conclusions The 10 min OCT and OCI were reliable predictors for the prognosis of patients with septic shock.
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Objective To investigate the clinical significance of soluble intercellular adhesion molecule-1 (sICAM-1) and interleukin-1β(IL-1β) in pregnant women with chorioamnionitis. Methods The levels of sICAM-1, IL-1β,CRP and WBC were detected by enzyme linked immunosorbent assay in maternal serum of 56 pregnant women with PPROM and 38 pregnant women with PROM. Every embryolemma was histopathologically confirmed after deliver-y. Results Chorioamnionitis in PPROM was more than that in PROM of full term(P<0.05). The serum levels of sI-CAM-1, IL-1β, CRP and WBC in chorioamnionitis in pregnant women were higher than those in non-chorioamnionitis (P <0.05). The serum levels of sICAM-1 was more sensitive and specific than those of CRP and WBC. The serum levels of IL-1β was more sensitive than those of CRP and WBC, but their specificity was similar. Conclusion The levels of sICAM-1 and IL-1β in maternal serum are good clinical biological markers for predicting chorioamnionitis in pregnant women.