Background: Preterm premature rupture of membranes remains a major complication after fetoscopic laser surgery (FLS) for twin-twin transfusion syndrome (TTTS). We studied the histologic changes of fetal membranes post-FLS and investigated a possible impact of amniotic fluid (AF) dilution. Methods: Fetal membranes of 31 pregnancies that underwent FLS for TTTS were investigated histologically at delivery at different sites: trocar site of recipient sac and at distance, donor sac, and inter-twin membrane. results: The trocar insertion site on the recipient sac showed no signs of histologic hallmarks of healing. Wide-spread alteration in collagen organization and higher apoptotic index in the amnion of the recipient sac which were absent in donor's and reference membranes. To explain the mechanisms, we analyzed the AF composition of recipient sacs from TTTS pregnancies vs. GA-matched healthy singleton controls and found glucose, protein and lactate dehydrogenase activity were all significantly lower in TTTS sacs consistent with over-dilution of recipient's AF (~2-fold). In-vitro exposure of healthy amniochorion to analogous dilutional stress conditions recapitulated the histologic changes and induced apoptosis and autophagy. conclusion: Alteration in structural integrity of the recipient's amniochorion, possibly in response to dilution stress, along with ineffective repair mechanisms may explain the increased incidence of preterm birth post-FLS.d espite the success of fetoscopic laser surgery (FLS) for the treatment of severe twin-twin transfusion (TTTS), preterm delivery with or without preterm premature rupture of membranes (PPROM) remains a major complication (1-4). The average gestational age at delivery after procedure ranges between 29 to 33 wk (1,5). Aside from the cost of caring for the premature infants, preterm delivery is directly associated with long-term neurological deficits in TTTS (4,6), undermining the complete benefits of such intervention. PPROM which affects 20-32% of cases is the most important risk factor leading to preterm delivery (2,7-10).Considered a minimally invasive procedure, FLS necessitates the insertion of a trocar into the amniotic cavity of the recipient twin to allow for insertion of the fetoscope (through a cannula which stays in place throughout the procedure) (11). A laser is used to seal the abnormal blood vessel anastomoses between the monochorionic twin placentas. Lastly, the surgeon performs an amnioreduction to relieve the recipient's polyhydramnios by draining the excess amniotic fluid (AF) through the cannula before removing it. During the procedure to allow proper visualization due to the inadvertent loss of amniotic fluid through cannula port or due to cloudy AF, it is customary for the surgeon to remove a volume of AF and replace it via amnioinfusion with a solution of Lactated Ringer's (LR).Fetal membranes are the outermost layer of fetal compartment in apposition to maternal decidua. The amniotic layer of the fetal membranes is made up of a lining of cuboidal epithe...