Dehydrated Amnion Chorion Membrane versus standard of care for diabetic foot ulcers: a randomised controlled trial.

OBJECTIVE: Diabetic foot ulcers (DFUs) continue to challenge wound care practitioners. This prospective, multicentre, randomised controlled trial (RCT) evaluated the effectiveness of a dehydrated Amnion Chorion Membrane (dACM) (Organogenesis Inc., US) versus standard of care (SoC) alone in complex DFUs in a challenging patient population. METHOD: Subjects with a DFU extending into dermis, subcutaneous tissue, tendon, capsule, bone or joint were enrolled in a 12-week trial. They were allocated equally to two treatment groups: dACM (plus SoC); or SoC alone. The primary endpoint was frequency of wound closure determined by a Cox analysis that adjusted for duration and wound area. Kaplan-Meier analysis was used to determine median time to complete wound closure (CWC). RESULTS: The cohort comprised 218 patients, and these were split equally between the two treatment groups with 109 patients in each. A Cox analysis showed that the estimated frequency of wound closure for the dACM plus SoC group was statistically superior to the SoC alone group at week 4 (12% versus 8%), week 6 (22% versus 11%), week 8 (31% versus 21%), week 10 (42% versus 27%) and week 12 (50% versus 35%), respectively (p=0.04). The computed hazard ratio (1.48 (confidence interval: 0.95, 2.29) showed a 48% greater probability of wound closure in favour of the dACM group. Median time to wound closure for dACM-treated ulcers was 84 days compared to 'not achieved' in the SoC-treated group (i.e., ≥50% of SoC-treated DFUs failed to heal by week 12; p=0.04). CONCLUSION: In an adequately powered DFU RCT, dACM increased the frequency, decreased the median time, and improved the probability of CWC when compared with SoC alone. dACM demonstrated beneficial effects in DFUs in a complex patient population. DECLARATION OF INTEREST: This study was funded by Organogenesis Inc., US. JC serves as a consultant and speaker for Organogenesis. RDD serves as a speaker for Organogenesis. OMA and MLS serve as consultants for Organogenesis. The authors have no other conflicts of interest to declare.

Orthologs of NOX5 and EC-SOD/SOD3: dNox and dSod3 Impact Egg Hardening Process and Egg Laying in Reproductive Function of Drosophila melanogaster.

The occurrence of ovarian dysfunction is often due to the imbalance between the formation of reactive oxygen species (ROS) and the ineffectiveness of the antioxidative defense mechanisms. Primary sources of ROS are respiratory electron transfer and the activity of NADPH oxidases (NOX) while superoxide dismutases (SOD) are the main key regulators that control the levels of ROS and reactive nitrogen species intra- and extracellularly. Because of their central role SODs are the subject of research on human ovarian dysfunction but sample acquisition is low. The high degree of cellular and molecular similarity between Drosophila melanogaster ovaries and human ovaries provides this model organism with the best conditions for analyzing the role of ROS during ovarian function. In this study we clarify the localization of the ROS-producing enzyme dNox within the ovaries of Drosophila melanogaster and by a tissue-specific knockdown we show that dNox-derived ROS are involved in the chorion hardening process. Furthermore, we analyze the dSod3 localization and show that reduced activity of dSod3 impacts egg-laying behavior but not the chorion hardening process.

Progesterone inactivation in decidual stromal cells: A mechanism for inflammation-induced parturition.

The process of human parturition involves inflammation at the interface where fetal chorion trophoblast cells interact with maternal decidual stromal (DS) cells and maternal immune cells in the decidua (endometrium of pregnancy). This study tested the hypothesis that inflammation at the chorion-decidua interface (CDI) induces labor by negating the capacity for progesterone (P4) to block labor and that this is mediated by inactivation of P4 in DS cells by aldo-keto reductase family 1 member C1 (AKR1C1). In human, Rhesus macaque, and mouse CDI, AKR1C1 expression increased in association with term and preterm labor. In a human DS cell line and in explant cultures of term human fetal membranes containing the CDI, the prolabor inflammatory cytokine, interleukin-1ß (IL-1ß), and media conditioned by LPS-stimulated macrophages increased AKR1C1 expression and coordinately reduced nuclear P4 levels and P4 responsiveness. Loss of P4 responsiveness was overcome by inhibition of AKR1C1 activity, inhibition of AKR1C1 expression, and bypassing AKR1C1 activity with a P4 analog that is not metabolized by AKR1C1. Increased P4 activity in response to AKR1C1 inhibition was prevented by the P4 receptor antagonist RU486. Pharmacologic inhibition of AKR1C1 activity prevented parturition in a mouse model of inflammation-induced preterm parturition. The data suggest that inflammatory stimuli at the CDI drive labor by inducing AKR1C1-mediated P4 inactivation in DS cells and that inhibiting and/or bypassing of AKR1C1-mediated P4 inactivation is a plausible therapeutic strategy to mitigate the risk of inflammation-associated preterm birth.

The relationship between thyroid autoantibodies and X chromosome monosomy in the chorionic tissue of patients with missed miscarriage.

OBJECTIVE: The aim of this study was to investigate the relationship between thyroid autoantibodies (TGAb and TPOAb) and X chromosome monosomy in the chorionic tissue of patients with missed early miscarriage. METHODS: The baseline data, thyroid function, thyroid antibody and the chromosomes from the chorionic tissue of 228 patients with missed early miscarriage were examined. RESULTS: (1) Among the 228 patients, 121 had a normal chromosome number, and 107 had an abnormal chromosome number. The majority of them were autosomal trisomy, of which trisomy 16 (40.19%) was predominant. Sex chromosome monosomy (28.04%) was secondary. (2) Among the 228 patients, 208 patients in this study had normal thyroid function (including 134 cases of negative thyroid antibodies and 74 cases of positive thyroid antibodies alone); 6 patients had abnormal thyroid function (including 2 cases of clinical hyperthyroidism, 3 cases of subclinical hypothyroidism, 1 case of hypothyroxinemia); and 14 patients had normal TSH and elevated T4 alone.(3) After exclusion of patients with thyroid function abnormalities, there were no significant differences in baseline data between the normal chromosome group and the abnormal chromosome group (P > 0.05). However, there was a significant difference in TGAb and TPOAb between the normal chromosome and abnormal chromosome group with 45, X karyotype, with a higher proportion of TGAb and/or TPOAb positivity in the 45, X karyotype group (P < 0.05). Additionally, compared to TGAb and/or TPOAb-positive patients, the risk of X chromosome monosomy was significantly reduced in TGAb and TPOAb-negative patients (P < 0.05). Moreover, both TGAb and TPOAb titer values in the X chromosome monosomy group were higher than those in the chromosomally normal group (P < 0.05). CONCLUSION: There is a correlation between TGAb, TPOAb and X chromosome monosomy in the chorionic tissue of patients with missed early miscarriage, although the mechanism remains to be further investigated.

Perinatal outcomes and growth discordance of triplet pregnancies based on chorionicity: a retrospective cohort study.

BACKGROUND: The worldwide occurrence of triplet pregnancy is estimated to be 0.093%, with a natural incidence of approximately 1 in 8000. This study aims to analyze the neonatal health status and birth weight discordance (BWD) of triplets based on chorionicity from birth until discharge. METHODS: This was a retrospective study. We reviewed a total of 136 triplet pregnancies at our tertiary hospital between January 1, 2001, and December 31, 2021. Maternal and neonatal outcomes, inter-triplet BWD, neonatal morbidity, and mortality were analyzed. RESULTS: Among all cases, the rates of intrauterine death, neonatal death, and perinatal death were 10.29, 13.07, and 24.26%, respectively. Thirty-seven of the cases resulted in fetal loss, including 13 with fetal anomalies. The maternal complications and neonatal outcomes of the 99 triplet pregnancies without fetal loss were compared across different chorionicities, including a dichorionic (DC) group (41 cases), trichorionic (TC) group (37 cases), and monochorionic (MC) group (21 cases). Neonatal hypoproteinemia (P < 0.001), hyperbilirubinemia (P < 0.019), and anemia (P < 0.003) exhibited significant differences according to chorionicity, as did the distribution of BWD (P < 0.001). More than half of the cases in the DC and TC groups had a BWD < 15%, while those in the MC group had a BWD < 50% (47.6%). TC pregnancy decreased the risk of neonatal anemia (adjusted odds ratio [AOR] = 0.084) and need for blood transfusion therapy after birth (AOR = 0.119). In contrast, a BWD > 25% increased the risk of neonatal anemia (AOR = 10.135) and need for blood transfusion after birth (AOR = 7.127). TC pregnancy, MCDA or MCTA, and BWD > 25% increased neonatal hypoproteinemia, with AORs of 4.629, 5.123, and 5.343, respectively. CONCLUSIONS: The BWD differed significantly according to chorionicity. Additionally, TC pregnancies reduced the risk of neonatal anemia and need for blood transfusion, but increased the risk of neonatal hypoproteinemia. In contrast, the BWD between the largest and smallest triplets increased the risk of neonatal anemia and the need for blood transfusion. TC pregnancy, MCDA or MCTA, and BWD > 25% increased the risks of neonatal hypoproteinemia. However, due to the limited number of triplet pregnancies, further exploration of the underlying mechanism is warranted.

Pregnancy outcomes of monochorionic diamniotic and dichorionic diamniotic twin pregnancies conceived by assisted reproductive technology and conceived naturally: a study based on chorionic comparison.

OBJECTIVE: To evaluate monochorionic diamniotic (MCDA) and dichorionic diamniotic (DCDA) twin pregnancies conceived by assisted reproductive technology (ART) and conceived naturally. METHODS: We retrospectively analyzed the data on twin pregnancies conceived by ART from January 2015 to January 2022,and compared pregnancy outcomes of MCDA and DCDA twins conceived by ART with those of MCDA and DCDA twins conceived naturally, pregnancy outcomes between MCDA and DCDA twins conceived by ART, and pregnancy outcomes of DCT and TCT pregnancies reduced to DCDA pregnancies with those of DCDA pregnancies conceived naturally. RESULT: MCDA pregnancies conceived by ART accounted for 4.21% of the total pregnancies conceived by ART and 43.81% of the total MCDA pregnancies. DCDA pregnancies conceived by ART accounted for 95.79% of the total pregnancies conceived by ART and 93.26% of the total DCDA pregnancies. Women with MCDA pregnancies conceived by ART had a higher premature delivery rate, lower neonatal weights, a higher placenta previa rate, and a lower twin survival rate than those with MCDA pregnancies conceived naturally (all p < 0.05). Women with DCDA pregnancies conceived naturally had lower rates of preterm birth, higher neonatal weights, and higher twin survival rates than women with DCDA pregnancies conceived by ART and those with DCT and TCT pregnancies reduced to DCDA pregnancies (all p < 0.05). CONCLUSION: Our study confirms that the pregnancy outcomes of MCDA pregnancies conceived by ART are worse than those of MCDA pregnancies conceived naturally. Similarly, the pregnancy outcomes of naturally-conceived DCDA pregnancies are better than those of DCDA pregnancies conceived by ART and DCT and TCT pregnancies reduced to DCDA pregnancies.

Monochorionic Twinning in Bioengineered Human Embryo Models.

Monochorionic twinning of human embryos increases the risk of complications during pregnancy. The rarity of such twinning events, combined with ethical constraints in human embryo research, makes investigating the mechanisms behind twinning practically infeasible. As a result, there is a significant knowledge gap regarding the origins and early phenotypic presentation of monochorionic twin embryos. In this study, a microthermoformed-based microwell screening platform is used to identify conditions that efficiently induce monochorionic twins in human stem cell-based blastocyst models, termed "twin blastoids". These twin blastoids contain a cystic GATA3+ trophectoderm-like epithelium encasing two distinct inner cell masses (ICMs). Morphological and morphokinetic analyses reveal that twinning occurs during the cavitation phase via splitting of the OCT4+ pluripotent core. Notably, each ICM in twin blastoids contains its own NR2F2+ polar trophectoderm-like region, ready for implantation. This is functionally tested in a microfluidic chip-based implantation assay with epithelial endometrium cells. Under defined flow regimes, twin blastoids show increased adhesion capacity compared to singleton blastoids, suggestive of increased implantation potential. In conclusion, the development of technology enabling large-scale formation of twin blastoids, coupled with high-sensitivity readout capabilities, presents an unprecedented opportunity for systematically exploring monochorionic twin formation and its impact on embryonic development.

PL-hMSC and CH-hMSC derived soluble factors inhibit proliferation but improve hGBM cell migration by activating TGF-ß and inhibiting Wnt signaling.

Glioblastoma multiforme (GBM) is one of the most common and aggressive brain tumors. GBM resists most chemotherapeutic agents, resulting in a high mortality rate in patients. Human mesenchymal stem cells (hMSCs), which are parts of the cancer stroma, have been shown to be involved in the development and progression of GBM. However, different sources of hMSCs might affect GBM cells differently. In the present study, we established hMSCs from placenta (PL-hMSC) and chorion (CH-hMSC) to study the effects of their released soluble factors on the proliferation, migration, invasion, gene expression, and survival of human GBM cells, U251. We found that the soluble factors derived from CH-hMSCs and PL-hMSCs suppressed the proliferation of U251 cells in a dose-dependent manner. In contrast, soluble factors derived from both hMSC sources increased U251 migration without affecting their invasive property. The soluble factors derived from these hMSCs decreased the expression levels of CyclinD1, E2Fs and MYC genes that promote GBM cell proliferation but increased the expression level of TWIST gene, which promotes EMT and GBM cell migration. The functional study suggests that both hMSCs might exert their effects, at least in part, by activating TGF-ß and suppressing Wnt/ß-catenin signaling in U251 cells. Our study provides a better understanding of the interaction between GBM cells and gestational tissue-derived hMSCs. This knowledge might be used to develop safer and more effective stem cell therapy that improves the survival and quality of life of patients with GBM by manipulating the interaction between hMSCs and GBM cells.

PURION® processed human amnion chorion membrane allografts retain material and biological properties supportive of soft tissue repair.

The reparative properties of amniotic membrane allografts are well-suited for a broad spectrum of specialties. Further enhancement of their utility can be achieved by designing to the needs of each application through the development of novel processing techniques and tissue configurations. As such, this study evaluated the material characteristics and biological properties of two PURION® processed amniotic membrane products, a lyophilized human amnion, intermediate layer, and chorion membrane (LHACM) and a dehydrated human amnion, chorion membrane (DHACM). LHACM is thicker; therefore, its handling properties are ideal for deep, soft tissue deficits; whereas DHACM is more similar to a film-like overlay and may be used for shallow defects or surgical on-lays. Characterization of the similarities and differences between LHACM and DHACM was conducted through a series of in vitro and in vivo studies relevant to the healing cascade. Compositional analysis was performed through histological staining along with assessment of barrier membrane properties through equilibrium dialysis. In vitro cellular response was assessed in fibroblasts and endothelial cells using cell proliferation, migration, and metabolic assays. The in vivo cellular response was assessed in an athymic nude mouse subcutaneous implantation model. The results indicated the PURION® process preserved the native membrane structure, nonviable cells and collagen distributed in the individual layers of both products. Although, LHACM is thicker than DHACM, a similar composition of growth factors, cytokines, chemokines and proteases is retained and consequently elicit comparable in vitro and in vivo cellular responses. In culture, both treatments behaved as potent mitogens, chemoattractants and stimulants, which translated to the promotion of cellular infiltration, neocollagen deposition and angiogenesis in a murine model. PURION® processed LHACM and DHACM differ in physical properties but possess similar in vitro and in vivo activities highlighting the impact of processing method on the versatility of clinical use of amniotic membrane allografts.

Dehydrated human amnion/chorion membrane to treat venous leg ulcers: a cost-effectiveness analysis.

OBJECTIVE: To evaluate the cost-effectiveness of dehydrated human amnion/chorion membrane (DHACM) in Medicare enrolees who developed a venous leg ulcer (VLU). METHOD: This economic evaluation used a four-state Markov model to simulate the disease progression of VLUs for patients receiving advanced treatment (AT) with DHACM or no advanced treatment (NAT) over a three-year time horizon from a US Medicare perspective. DHACM treatments were assessed when following parameters for use (FPFU), whereby applications were initiated 30-45 days after the initial VLU diagnosis claim, and reapplications occurred on a weekly to biweekly basis until completion of the treatment episode. The cohort was modelled on the claims of 530,220 Medicare enrolees who developed a VLU between 2015-2019. Direct medical costs, quality-adjusted life years (QALYs), and the net monetary benefit (NMB) at a willingness-to-pay threshold of $100,000/QALY were applied. Univariate and probabilistic sensitivity analyses (PSA) were performed to test the uncertainty of model results. RESULTS: DHACM applied FPFU dominated NAT, yielding a lower per-patient cost of $170 and an increase of 0.010 QALYs over three years. The resulting NMB was $1178 per patient in favour of DHACM FPFU over the same time horizon. The rate of VLU recurrence had a notable impact on model uncertainty. In the PSA, DHACM FPFU was cost-effective in 63.01% of simulations at the $100,000/QALY threshold. CONCLUSION: In this analysis, DHACM FPFU was the dominant strategy compared to NAT, as it was cost-saving and generated a greater number of QALYs over three years from the US Medicare perspective. A companion VLU Medicare outcomes analysis revealed that patients who received AT with a cellular, acellular and matrix-like product (CAMP) compared to patients who received NAT had the best outcomes. Given the added clinical benefits to patients at lower cost, providers should recommend DHACM FPFU to patients with VLU who qualify. Decision-makers for public insurers (e.g., Medicare and Medicaid) and commercial payers should establish preferential formulary placement for reimbursement of DHACM to reduce budget impact and improve the long-term health of their patient populations dealing with these chronic wounds. DECLARATION OF INTEREST: Support for this analysis was provided by MiMedx Group, Inc., US. JLD, and RAF are employees of MiMedx Group, Inc. WHT, BH, PS, BGC and WVP were consultants to MiMedx Group, Inc. VD, AO, MRK, JAN, NW and GAM served on the MiMedx Group, Inc. Advisory Board. MRK and JAN served on a speaker's bureau. WVP declares personal fees and equity holdings from Stage Analytics, US.

Evaluation of Regenerative Efficacy of Amnion and Chorion Membrane in Treatment of Mandibular Molar Furcation Defects: A Clinico-radiographic Study.

AIM: Amnion and chorion membranes possess unique inherited biological properties that enhance wound healing and may accelerate periodontal regeneration. The present study aims to evaluate and compare the efficacy of amnion and chorion membranes in the treatment of furcation defects. MATERIALS AND METHODS: A total of 20 patients were selected and were randomly allocated to group I and group II with 10 subjects in each group. Amnion and chorion membranes are placental-derived membranes that accelerate regeneration by having natural growth factors with their antimicrobial and inflammation reduction properties. Group I was treated using bone grafting with decalcified freeze-dried bone allograft (DFDBA) and placement of amnion as a membrane for guided tissue regeneration (GTR) whereas group II was treated using bone grafting with DFDBA and placement of chorion as a membrane for GTR. The patients were followed for clinical and radiographic parameters and were evaluated between 3 and 6 months after surgery. RESULT: In intragroup comparison, a significant difference was evident in both the groups for all the clinical and radiographic parameters within the groups. (p = 0.01) This means both amnion and chorion membranes showed statistically significant regenerative efficacy. In intergroup comparison, the results show that all the clinical parameters and radiographic parameters show no significant difference between the groups. CONCLUSION: The amnion and chorion membranes had similar regenerative efficacy in combination with DFDBA in patients with buccal degree II furcation defects in mandibular molars. CLINICAL SIGNIFICANCE: The amnion and chorion membranes have shown significant improvement in clinical and radiographic parameters when used for the treatment of buccal degree II furcation defects in mandibular molars. How to cite this article: Mallapragda S, Gupta R, Gupta S, et al. Evaluation of Regenerative Efficacy of Amnion and Chorion Membrane in Treatment of Mandibular Molar Furcation Defects: A Clinico-radiographic Study. J Contemp Dent Pract 2024;25(2):160-167.

Evaluation of Leukocyte Chemotaxis Induced by Human Fetal Membranes in an In Vitro Model.

Leukocyte infiltration into the maternal-fetal interface is a consequence of the robust inflammation in the gestational tissues during term labor and preterm labor with or without infection. During pregnancy, the fetal membranes act as a physical barrier that isolates the fetus into the amniotic cavity, keeping it in an optimal environment for its development. In addition, the fetal membranes possess immunological competencies such as the secretion of cytokines and chemokines in response to different stimuli. Clinical and experimental evidence indicates that these tissues are involved in the extensive chemotaxis of immune cells in normal or pathological conditions.Few studies have evaluated the chemotactic capacities of the fetal membranes considering that this tissue is composed of two adjacent tissues, the amnion and the chorion, which have different characteristics. Although these tissues function as a unit, their response is complex since there is an interaction between them, where each tissue contributes differently. The protocol described here allows us to evaluate the in vitro chemotactic capacities of fetal membranes in response to various applied stimuli, considering the contribution of each of their components (amnion and choriodecidua) using a Boyden chamber assay and phenotyping the chemo-attracted leukocytes by flow cytometry.

Culture of Human Fetal Membranes in a Two Independent Compartment Model: An Ex Vivo Approach.

During pregnancy, the fetal membranes composed of the amnion and chorodecidua constitute a selective barrier separating two distinct environments, maternal and fetal. These tissues have the function of delimiting the amniotic cavity. Their histological complexity gives them physical, mechanical, and immunological properties to protect the fetus. Although the study of the amnion, chorion, and decidua separately provides knowledge about the functions of the fetal membranes, the protocol we describe in this chapter has the advantage of maintaining the biological and functional complexity of these tissues. In addition, this experimental model allows the researcher to recreate various pathological scenarios because this model allows for differential stimulation of the amnion or choriodecidua.

Temporal ultrasound findings in the first trimester of a dual fetal demise occurring in a monochorionic diamniotic twin gestation.

Single fetal demise in monochorionic gestations in the 2nd and 3rd trimester is associated with adverse outcomes for the co-twin. We present a case of single demise in a monochorionic gestation in the 1st trimester with evidence of subsequent hemodynamic aberrations in the co-twin, supportive of feto-fetal hemorrhage occurring early in gestation.

Membrane inflammasome activation by choriodecidual Ureaplasma parvum infection without intra-amniotic infection in a Non-Human Primate model†.

Intrauterine infection is a significant cause of neonatal morbidity and mortality. Ureaplasma parvum is a microorganism commonly isolated from cases of preterm birth and preterm premature rupture of membranes (pPROM). However, the mechanisms of early stage ascending reproductive tract infection remain poorly understood. To examine inflammation in fetal (chorioamnionic) membranes we utilized a non-human primate (NHP) model of choriodecidual U. parvum infection. Eight chronically catheterized pregnant rhesus macaques underwent maternal-fetal catheterization surgery at ~105-112 days gestation and choriodecidual inoculation with U. parvum (105 CFU/mL, n =4) or sterile media (controls; n = 4) starting at 115-119 days, repeated at 5-day intervals until C-section at 136-140 days (term=167 days). The average inoculation to delivery interval was 21 days, and Ureaplasma infection of the amniotic fluid (AF) was undetectable in all animals. Choriodecidual Ureaplasma infection resulted in increased fetal membrane expression of MMP-9 and PTGS2, but did not result in preterm labor or increased concentrations of AF pro-inflammatory cytokines. However, membrane expression of inflammasome sensors, NLRP3, NLRC4, AIM2, and NOD2, and adaptor ASC (PYCARD) gene expression were significantly increased. Gene expression of IL-1ß, IL-18, IL-18R1  , CASPASE-1, and pro-CASPASE-1 protein increased with Ureaplasma infection. Downstream inflammatory genes MYD88 and NFκB (Nuclear factor kappa-light-chain-enhancer of activated B cells) were also significantly upregulated. These results demonstrate that choriodecidual Ureaplasma infection, can cause activation of inflammasome complexes and pathways associated with pPROM and preterm labor prior to microbes being detectable in the AF.

Preterm membranes are mechanically more resistant than term membranes.

OBJECTIVE: To compare the biomechanical properties of fetal preterm membranes (20 + 0 weeks to 30 + 0 weeks) to those of the term (37 + 0 to 41 + 0 weeks). METHOD: Amnion and chorion were manually separated and samples were cut to the required geometry. Rectangular samples with (mode 1) and without (uniaxial) a notch, were tested for tearing energy, critical elongation, and tangent stiffness. Suture retention and inter-suture distance testing investigated the effect of suture placement. RESULTS: From the 15 preterm and 10 term placentas studied, no notable differences were observed in uniaxial testing. Mode 1 fracture testing showed a difference in tearing energy between the preterm and term chorion (0.025 ± 0.005 vs. 0.017 ± 0.005 J/m-1 ; p = 0.027) but not in the amnion (0.030 ± 0.017 vs. 0.029 ± 0.009 J/m-1 ; p = 0.895). Both preterm amnion and chorion showed a higher critical elongation compared with term (1.229 ± 0.057 vs. 1.166 ± 0.046; p = 0.019 and 1.307 ± 0.049 vs. 1.218 ± 0.058; p = 0.012). Preterm amnion had a higher suture retention strength than its term counterpart (0.189 ± 0.065 vs. 0.121 ± 0.031 N; p = 0.023). In inter-suture distance tests, no significant interaction was observed beyond 3 mm, but the preterm chorion showed less interaction at 1-2 mm distances. CONCLUSION: Preterm membranes have equivalent or superior tensile properties to term membranes. The chorion appears to contribute to the mechanical integrity of fetal membranes, particularly in preterm stages.

Zebrafish prss59.1 is involved in chorion development.

The prss59.1 gene was identified as one of 11 genes that were highly upregulated during the induction of ovulation in zebrafish by using an in vivo ovulation assay. Previously, we conducted biochemical characterization of Prss59.1 and revealed it to be a trypsin-like proteolytic enzyme. In this study, we established a prss59.1 gene knockout strain using the CRISPR/Cas9 system. Phenotypic analysis of prss59.1 knockout fish showed that prss59.1 is associated with chorion elevation, a prominent event in egg activation during fertilization. The chorions of heterozygous and homozygous prss59.1 mutant zebrafish were smaller than those of the wild type. The results suggested that Prss59.1 is necessary for chorion expansion. The homozygous prss59.1 mutant strain, with a small chorion, showed an extremely low survival rate. Fiber-supported knob-like structures (KS) on the chorion showed an abnormal structure in prss59.1 mutants. Prss59.1 was detected in the KS on the chorion. The pores on the chorion were smaller in the prss59.1 mutants than in the wild type. Transmission electron microscopy (TEM) observations of the cross sections of the chorions showed abnormalities in the chorion structure in prss59.1 mutants. These results demonstrated that Prss59.1 is involved in chorion elevation and in proper formation of the chorion, which is necessary for embryo development.

Anatomy of the fetal membranes: insights from spinning disk confocal microscopy.

PURPOSE: The fetal membranes are essential for the maintenance of pregnancy, and their integrity until parturition is critical for both fetal and maternal health. Preterm premature rupture of the membranes (pPROM) is known to be an indicator of preterm birth, but the underlying architectural and mechanical changes that lead to fetal membrane failure are not yet fully understood. The aim of this study was to gain new insights into the anatomy of the fetal membrane and to establish a tissue processing and staining protocol suitable for future prospective cohort studies. METHODS: In this proof of principle study, we collected fetal membranes from women undergoing vaginal delivery or cesarean section. Small membrane sections were then fixed, stained for nucleic acids, actin, and collagen using fluorescent probes, and subsequently imaged in three dimensions using a spinning disk confocal microscope. RESULTS: Four fetal membranes of different types were successfully processed and imaged after establishing a suitable protocol. Cellular and nuclear outlines are clearly visible in all cases, especially in the uppermost membrane layer. Focal membrane (micro) fractures could be identified in several samples. CONCLUSION: The presented method proves to be well suited to determine whether and how the occurrence of membrane (micro) fractures and cellular jamming correlate with the timing of membrane rupture and the mode of delivery. In future measurements, this method could be combined with mechanical probing techniques to compare optical and mechanical sample information.