Prediction And Prevention Of Ovarian Hyperstimulation Syndrome (OHSS) And The Role Of Low Molecular Weight Heparin (LMWH) In OHSS Prevention And Management

OHSS is a dramatic complication of ovulation induction. Ovulation accompanied by tissue damage can cause an increase in both plasma and follicular fluid (FF) level of both tissue factor (TF) and vascular endothelial growth factor (VEGF) and decrease in tissue factor pathway inhibitor (TFPI) level especially with controlled ovarian hyperstimulation (COH) while undergoing In vitro fertilization (ⅣF).Materials&methods:A prospective cohort study with matched controls of293patients were divided into3groups, group Ⅰ; PCO or PCOS patients (randomly divided into2subgroups; IA and ⅠB with their matched controls; ⅠC), group Ⅱ; patients at high risk of developing OHSS (randomly divided into2subgroups; ⅡA and IIB with their matched controls;ⅡC), group III; severe OHSS patients (randomly divided into2subgroups; ⅢA and ⅢB), group IA patients were injected by Fraxiparine (速碧林注射剂)4100IU once daily began with the start of COH while in group IIA it was started on HCG day until pregnancy was confirmed or not. Group IIIA patients, as well, were injected by Fraxiparine once OHSS developed until recovery. Groups IB, ⅡB and IIIB were injected by placebo on the same time points. Plasma and FF concentrations of VEGF,TF,TFPI on successive4time points{on gonadotropin start, HCG day, ovum retrieval (ORJday, embryo transfer (ET)day) were detected using ELISA kits in group Ⅰ and Ⅱ, while plasma concentrations of the same factors were detected every other day in OHSS patients(group Ⅲ).In vitro, granulose cells (GC) of60patients (PCO/PCOS; group Ⅰ, patients at risk of developing OHSS; group Ⅱ and matched controls) were collected (20from each group) and primary cultured with human chorionic gonadotropin (hCG,1IU/ml) and with adding different concentrations of Fraxiparine (0,5,50μg/ml) in the culture media for3days duration. Granulosa cells were harvested and mRNA was extracted and cDNA was used with primers of VEGF165, VEGF121, TF, TFPI, and GAPDH in real time PCR to detect their expression levels.Moreover, Fraxiparine therapeutic effect was checked in OHSS patients and monitored by plasma levels of the same three factors. Also, the mechanism by which Fraxiparine affect (GC) was checked by primary culture of (GC) with adding Fraxiparine and inhibitors of three signal transduction pathways (NF-kB, JNK, Akt) with repeating the real time PCR for detecting the expression level of VEGF165, VEGF121, TF, TFPI and GAPDHIn addition, VEGF receptors (VEGF R1and VEGF R2) in ovarian tissue samples had been checked using immunoflourescence. Also, both receptors in primary cultured granulosa cells had been checked using Immunoflourescence. Ovarian tissue samples were obtained from both normal control patients and PCO/PCOS patients. Granulosa cells were obtained during Egg Retrieval from normal control, PCO/PCOS and patients at high risk of developing OHSS (either with high E2level and with multiple mature follicles>20) and cultured for48hours.Finally, Western Blot was done for detecting the protein level of both receptors in granulosa cells obtained from the same three groups of infertile patients.RESULTS:When comparing groups IB and IIB, plasma and FF levels of VEGF and TF were significantly higher while TFPI levels was significantly lower than their comparable levels in matched controls on all time points except TFPI on Egg Retrieval day and FF samples when comparing group IB with their controls. Same groups showed significantly higher mRNA expression levels of VEGF165, VEGF121, TF, and significantly lower mRNA expression level of TFPI than in their controls except with VEGF121and TFPI levels when comparing group I and II with their controls respectively. ELISA showed significantly higher levels of VEGF and TF and significantly lower level of TFPI in culture media of the same groups.On all time points, Fraxiparine decreased the plasma and FF levels of VEGF and TF in group IA significantly,also, it increased the plasma and FF levels of TFPI significantly than those in group IB. Developing OHSS in group IA was significantly lower than that in group IB and IIB. Same groups showed significantly lower mRNA expression levels of VEGF165, VEGF121, TF, and significantly higher mRNA expression level of TFPI than in their controls. ELISA showed significantly Lower levels of VEGF and TF and significantly higher level of TFPI in culture media of same groups.Both early and late OHSS patients in group ⅢA showed significantly lower plasma levels of VEGF, TF and significantly higher levels of TFPI than those in group IIIB.In group Ⅰ and Ⅱ granulose cells, Fraxiparine effect was inhibited by cellular transduction pathway (NF-kB, JNK, Akt) inhibitors. Immunoflourescence showed that VEGF receptors (VEGF R1and VEGF R2) were expressed more in PCO/PCOS patients than their expression in normal control patients. Immunoflourescence and Western Blot showed that VEGF receptors (VEGF R1and VEGF R2) and their protein levels were expressed more in both PCO/PCOS group AND patients at high risk of developing OHSS than their expression in normal control patients.CONCLUSIONS:VEGF, TF and TFPI can predict the occurrence of OHSS in both PCO/PCOS infertile patients and those at high risk of developing OHSS. Fraxiparine, if used with the start of controlled ovarian hyperstimulation, can prevent the developing of OHSS and manage OHSS if already developed in those infertile patients.