Implantation (embryology)

From Wikipedia, the free encyclopedia
  (Redirected from Implantation (human embryo))
Jump to navigation Jump to search
The sperm and ovum unite through fertilization, creating an embryo that over the course of 8–9 days in the human will implant in the uterine wall, where it will reside over the nine-month course of gestation.

In female mammals implantation (also known as nidation) is the stage in embryonic development in which the blastocyst hatches as the embryo, and adheres to the wall of the uterus.[1] Once this adhesion is successful, the female is considered to be pregnant and the embryo will receive oxygen and nutrients from the mother in order to grow.

There is an extensive variation in the type of trophoblast cells, and structures of the placenta across the different species of mammals.[2] However, of the five recognised stages of implantation preceding placentation the first four are similar across the species. The five stages are migration and hatching, pre-contact, attachment, adhesion, and invasion.[2] Some sources only recognize the last three stages.[3][4]

In humans implantation of a fertilized ovum usually takes place around nine days after ovulation but can range between six and twelve days.[5]

Uterus receptivity[edit]

To enable implantation, the uterus goes through changes in order to be able to receive the conceptus. Receptivity includes changes to endometrial cells that help to absorb uterine fluid; changes in the thickness of the endometrium and its blood supply development, and the formation of the decidua. Collectively these changes are known as plasma membrane transformation, and bring the blastocyst nearer to the endometrium and immobilize it. During this stage the blastocyst can still be eliminated by being flushed out of the uterus.

Successful implantation is co-dependent on the viability of the embryo, and the receptivity of the uterus.[6] A critical involved factor is the developmental synchrony between the embryo and the uterus.[7] The synchrony gives a short period of receptivity known as the implantation window.[8][9]

In humans uterine receptivity is optimum on days 20-24 of the secretory phase of the menstrual cycle when luteinizing hormone levels are at their peak.[3][10][6] The endometrial microbiome has been indicated as having an important role in successful implantation in controlling endometrial cell function, and the function of the local immunity system that prevents pathogen growth. This is associated with the secretion of protective substances.[11][12]

Implantation stages[edit]

There are five recognized stages of implantation in mammals that precede the formation of the placenta. They are: migration and hatching, pre-contact, attachment, adhesion, and invasion. The first four stages are similar across the species with process of invasion being variable.[2] Many sources only recognize the last three three stages.[13] These three stages of apposition, attachment, and invasion are also alternatively termed contact (apposition), adhesion (attachment), and penetration (invasion).[4][3]

Predecidualization[edit]

The endometrium increases thickness, becomes vascularized and its glands grow to be tortuous and boosted in their secretions. These changes reach their maximum about seven days after ovulation.

Furthermore, the surface of the endometrium produces a kind of rounded cells, which cover the whole area toward the uterine cavity. This happens about 9 to 10 days after ovulation.[14] These cells are called decidual cells, which emphasises that the whole layer of them is shed off in every menstruation if no pregnancy occurs, just as leaves of deciduous trees. The uterine glands, on the other hand, decrease in activity and degenerate around 8 to 9 days[14] after ovulation in absence of pregnancy.

The decidual cells originate from the stromal cells that are always present in the endometrium, and make up a new layer, the decidua. The rest of the endometrium, in addition, expresses differences between the luminal and the basal sides. The luminal cells form the stratum compactum of the endometrium, in contrast to the basalolateral stratum spongiosum, which consists of the rather spongy stromal cells.[14]

Decidualization[edit]

Decidualization succeeds predecidualization if pregnancy occurs. This is an expansion of it, further developing the uterine glands, the zona compacta and the epithelium of decidual cells lining it. The decidual cells become filled with lipids and glycogen and take the polyhedral shape characteristic of decidual cells.

Trigger[edit]

It is likely that the blastocyst itself makes the main contribution to this additional growing and sustaining of the decidua. An indication of this is that decidualization occurs at a higher degree in conception cycles than in nonconception cycles.[14] Furthermore, similar changes are observed when giving stimuli mimicking the natural invasion of the embryo.[14]

The embryo releases serine proteases which causes the epithelial cell membrane to depolarize and activates the epithelial Na+ channel. This triggers a Ca2+ influx and phosphorylation of CREB. Phosphorylation of CREB upregulates the expression of COX-2, which leads to the release of prostaglandin E2 (PGE2) from epithelial cells. PGE2 acts on the stroma cells activating cAMP-related pathways in stromal cell leading to decidualization.[15][16]

Parts of decidua[edit]

The decidua can be organized into separate sections, although they have the same composition.

  • Decidua basalis – This is the part of the decidua which is located basalolateral to the embryo after implantation.
  • Decidua capsularis – Decidua capsularis grows over the embryo on the luminal side, enclosing it into the endometrium. It surrounds the embryo together with decidua basalis.
  • Decidua parietalis – All other decidua on the uterine surface belongs to decidua parietalis.

Decidua throughout pregnancy[edit]

After implantation the decidua remains, at least through the first trimester.[14] However, its most prominent time is during the early stages of pregnancy, during implantation. Its function as a surrounding tissue is replaced by the definitive placenta. However, some elements of the decidualization remain throughout pregnancy.[14]

The compacta and spongiosa layers are still observable beneath the decidua in pregnancy. The glands of the spongiosa layer continue to secrete during the first trimester, when they degenerate. However, before that disappearance, some glands secrete unequally much. This phenomenon of hypersecretion is called the Arias-Stella phenomenon,[14] after the pathologist Javier Arias-Stella.

Pinopodes[edit]

Pinopodes are small, finger-like protrusions from the endometrium. They appear between day 19 and day 21 of gestational age.[14] This corresponds to a fertilization age of approximately five to seven days, which corresponds well with the time of implantation. Pinopodes only persist for two to three days.[14] Their development is enhanced by progesterone, and inhibited by estrogens. Pinopodes endocytose uterine fluid and its macromolecules. By doing so, the volume of the uterus decreases, taking the walls closer to the embryoblast floating in it. Thus, the period of active pinopodes might also limit the implantation window.[14] During the early stages of implatation pinopodes continue to absorb fluid, and remove most of it during the early stages of implantation.

Adaptation of secretions[edit]

Proteins, glycoproteins and peptides

secreted by the endometrial glands[14]

Matrix-associated:
Fibronectin
Laminin
Entactin
Type-IV collagen
heparan sulfate
Proteoglycan
Integrins
Others:
Mucins
Prolactin
IGFBP-1
Glycodelin
Pregnancy-associated endometrial

alpha-2-globulin (alpha-2-PEG)

endometrial protein 15
Albumin
Beta-Lipoprotein
Relaxin
Fibroblast growth factor 1
Fibroblast growth factor 2
Pappalysin-1
Stress response protein 27 (SRP-27)
CA-125
Beta-endorphin
Leu-enkephalin
Diamine oxidase
Tissue plasminogen activator
Renin
Progesterone-dependent carbonic anhydrase
Lactoferrin

Not only the lining of the uterus transforms, but in addition, the secretion from its epithelial glands changes. This change is induced by increased levels of progesterone from the corpus luteum. The target of the secretions is the embryoblast, and has several functions on it.

Nourishment[edit]

The embryoblast spends approximately 72 hours in the uterine cavity before implanting. In that time, it cannot receive nourishment directly from the blood of the mother, and must rely on secreted nutrients into the uterine cavity, e.g. iron and fat-soluble vitamins.[14]

Growth and implantation[edit]

In addition to nourishment, the endometrium secretes several steroid-dependent proteins, important for growth and implantation. Cholesterol, and steroids are also secreted.[14] Implantation is further facilitated by synthesis of matrix substances, adhesion molecules and surface receptors for the matrix substances.

Mechanism[edit]

Zona hatching[edit]

Implantation is initiated when the blastocyst hatches as the embryo, and comes into contact with the uterine wall. Hatching involves the ridding of the zona pellucida.

Lytic factors in the uterine cavity, as well as factors from the blastocyst itself are essential for this process. Mechanisms in the latter are indicated by that the zona pellucida remains intact if an unfertilized egg is placed in the uterus under the same conditions.[14] A substance probably involved is plasmin. Plasminogen, the plasmin precursor, is found in the uterine cavity, and blastocyst factors contribute to its conversion to active plasmin. This hypothesis is supported by lytic effects in vitro by plasmin.[14] Furthermore, plasmin inhibitors also inhibit the entire zona hatching in rat experiments.[14]

Apposition[edit]

The very first, loose connection between the blastocyst and the endometrium is called the apposition.[14]

Location[edit]

On the endometrium, the apposition is usually made where there is a small crypt in it, perhaps because it increases the area of contact with the rather spherical blastocyst.

On the blastocyst, on the other hand, it occurs at a location where there has been enough lysis of the zona pellucida to have created a rupture to enable direct contact between the underlying trophoblast and the decidua of the endometrium.[14] However, ultimately, the inner cell mass, inside the trophoblast layer, is aligned closest to the decidua. Nevertheless, the apposition on the blastocyst is not dependent on if it is on the same side of the blastocyst as the inner cell mass. Rather, the inner cell mass rotates inside the trophoblast to align to the apposition.[14] In short, the entire surface of the blastocyst has a potential to form the apposition to the decidua.

Molecular mechanism[edit]

The identity of the molecules on the trophoblast and the endometrial epithelia that mediate the initial interaction between the two remain unidentified. However, a number of research groups have proposed that MUC1, a member of the mucin family of glycosylated proteins, is involved.[17] MUC1 is a transmembrane glycoprotein expressed at the apical surface of endometrial epithelial cells during the window of implantation in humans and has been shown to be differentially expressed between fertile and infertile subjects during this time.[17] MUC1 displays carbohydrate moieties on its extracellular domain that are ligands of L-selectin, a protein expressed on the surface of trophoblast cells.[18] An in vitro model of implantation developed by Genbacev et al., gave evidence to support the hypothesis that L-selectin mediates apposition of the blastocyst to the uterine epithelium by interacting with its ligands.[19]

Adhesion[edit]

Adhesion is a much stronger attachment to the endometrium than the loose apposition.

The trophoblasts adhere by penetrating the endometrium, with protrusions of trophoblast cells.

This adhering activity is by microvilli that are on the trophoblast. The trophoblast have binding fiber connections, laminin, collagen type IV, and integrins that assist in this adhesion process.[20]

MUC16 is a transmembrane mucin expressed at the apical surface of uterine epithelia. This mucin prevents the blastocyst from implanting in an undesired located on the epithelium. Thus, MUC16 inhibits cell-cell adhesion. "Removal of this mucin during formation of uterodomes (bulbous projections from the apical surface of the epithelium that are often found during the implantation period) facilitates trophoblast adhesion in vitro".[21]

Communication[edit]

There is massive communication between the blastocyst and the endometrium at this stage. The blastocyst signals to the endometrium to adapt further to its presence, e.g. by changes in the cytoskeleton of decidual cells. This, in turn, dislodges the decidual cells from their connection to the underlying basal lamina, which enables the blastocyst to perform the succeeding invasion.[14]

This communication is conveyed by receptor-ligand-interactions, both integrin-matrix and proteoglycan ones.

Proteoglycan receptors[edit]

Another ligand-receptor system involved in adhesion is proteoglycan receptors, found on the surface of the decidua of the uterus. Their counterparts, the proteoglycans, are found around the trophoblast cells of the blastocyst. This ligand-receptor system also is present just at the implantation window.[14]

Invasion[edit]

Invasion is an even further establishment of the blastocyst in the endometrium.

Syncytiotrophoblasts[edit]

The protrusions of trophoblast cells that adhere into the endometrium continue to proliferate and penetrate into the endometrium. As these trophoblast cells penetrate, they differentiate to become a new type of cells, syncytiotrophoblast. The prefix syn- refers to the transformation that occurs as the boundaries between these cells disappear to form a single mass of many cell nuclei (a syncytium). The rest of the trophoblasts, surrounding the inner cell mass, are hereafter called cytotrophoblasts. Syncytiotrophoblast is not determined as a cell type, rather is a multinucleated tissue [22]

Invasion continues with the syncytiotrophoblasts reaching the basal membrane beneath the decidual cells, penetrating it and further invading into the uterine stroma. Finally, the whole embryo is embedded in the endometrium. Eventually, the syncytiotrophoblasts come into contact with maternal blood and form chorionic villi. This is the initiation of forming the placenta.

The penetration of the trophoblast to the endometrium is demonstrated through metalloproteinase MMP-2 and MMP-9.[23] Syncytiotrophoblast invade the uterus attempting to reach maternal blood supply, for setting up the foundation for fetal blood flow [24]

Extravillous trophoblasts[edit]

Extravillous trophoblasts are cells from the invading villi that migrate into the myometrium of the mother’s uterus. These cells remodel the spiral arteries to improve and secure maternal blood flow to the growing embryo. There is also evidence that this process occurs with the uterine veins, stabilizing them to improve drainage of fetal blood and metabolic wastes.[25] Trophoblasts have also been documented to migrate into the mother and have been found in various tissues. Due to this trophoblasts have been implicated in a phenomenon known as "Fetomaternal Microchimerism" where fetal cells establish cell lines in maternal tissues.[26]

Secretions[edit]

The blastocyst secretes factors for a multitude of purposes during invasion. It secretes several autocrine factors, targeting itself and stimulating it to further invade the endometrium.[14] Furthermore, secretions loosen decidual cells from each other, prevent the embryo from being rejected by the mother, trigger the final decidualization and prevent menstruation.

Autocrine[edit]

Human chorionic gonadotropin is an autocrine growth factor for the blastocyst.[14] Insulin-like growth factor 2,[14] on the other hand, stimulates the invasiveness of it.

Dislodging[edit]

The syncytiotrophoblasts dislodges decidual cells in their way, both by degradation of cell adhesion molecules linking the decidual cells together as well as degradation of the extracellular matrix between them.

Cell adhesion molecules are degraded by syncytiotrophoblast secretion of tumor necrosis factor-alpha. This inhibits the expression of cadherins and beta-catenin.[14] Cadherins are cell adhesion molecules, and beta-catenin helps to anchor them to the cell membrane. Inhibited expression of these molecules thus loosens the connection between decidual cells, permitting the syncytotrophoblasts and the whole embryo with them to invade into the endometrium.

The extracellular matrix is degraded by serine endopeptidases and metalloproteinases. Examples of such metalloproteinases are collagenases, gelatinases and stromelysins.[14] These collagenases digest Type-I collagen, Type-II collagen, Type-III collagen, Type-VII collagen and Type-X collagen.[14] The gelatinases exist in two forms; one digesting Type-IV collagen and one digesting gelatin.[14]

Immunosuppressive[edit]

The embryo differs from the cells of the mother, and would be rejected as a parasite by the immune system of the mother if it did not secrete immunosuppressive agents. Such agents include platelet-activating factor, human chorionic gonadotropin, early pregnancy factor, prostaglandin E2, interleukin 1-alpha, interleukin 6, interferon-alpha, leukemia inhibitory factor and colony-stimulating factor.

Decidualization[edit]

Factors from the blastocyst also trigger the final formation of decidual cells into their proper form. In contrast, some decidual cells in the proximity of the blastocyst degenerate, providing nutrients for it.[14]

Prevention of menstruation[edit]

Human chorionic gonadotropin (hCG) not only acts as an immunosuppressive,[14] but also "notifies" the mother's body that she is pregnant, preventing menstruation by sustaining the function of the corpus luteum.

Other factors[edit]

Other factors secreted by the blastocyst are;

Failure[edit]

Implantation failure is considered to be caused by inadequate uterine receptivity in two-thirds of cases, and by problems with the embryo itself in the other third.[28]

Inadequate uterine receptivity may be caused by abnormal cytokine and hormonal signaling as well as epigenetic alterations.[29] Recurrent implantation failure is a cause of female infertility. Therefore, pregnancy rates can be improved by optimizing endometrial receptivity for implantation.[29] Evaluation of implantation markers may help to predict pregnancy outcome and detect occult implantation deficiency.[29]

Luteal support is the administration of medication, generally progestins, for the purpose of increasing the success rate of implantation and early embryogenesis, thereby complementing the function of the corpus luteum.

In women with more than three implantation failures in assisted reproduction, a review of several small randomized controlled studies estimated that the use of adjunct low molecular weight heparin (LMWH) improves live birth rate by approximately 80%.[30]

See also[edit]

References[edit]

  1. ^ Niringiyumukiza, JD; Cai, H; Xiang, W (1 May 2018). "Prostaglandin E2 involvement in mammalian female fertility: ovulation, fertilization, embryo development and early implantation". Reproductive biology and endocrinology : RB&E. 16 (1): 43. doi:10.1186/s12958-018-0359-5. PMID 29716588.
  2. ^ a b c Imakawa, K; Bai, R; Fujiwara, H; Kusama, K (January 2016). "Conceptus implantation and placentation: molecules related to epithelial-mesenchymal transition, lymphocyte homing, endogenous retroviruses, and exosomes". Reproductive medicine and biology. 15 (1): 1–11. doi:10.1007/s12522-015-0215-7. PMID 29259417.
  3. ^ a b c Kim, SM; Kim, JS (December 2017). "A Review of Mechanisms of Implantation". Development & reproduction. 21 (4): 351–359. doi:10.12717/DR.2017.21.4.351. PMID 29359200.
  4. ^ a b McGowen, MR; Erez, O; Romero, R; Wildman, DE (2014). "The evolution of embryo implantation". The International journal of developmental biology. 58 (2–4): 155–61. doi:10.1387/ijdb.140020dw. PMID 25023681.
  5. ^ Wilcox AJ, Baird DD, Weinberg CR (1999). "Time of implantation of the Conceptus and loss of pregnancy". New England Journal of Medicine. 340 (23): 1796–1799. doi:10.1056/NEJM199906103402304. PMID 10362823.
  6. ^ a b Yang, Y; Zhu, QY; Liu, JL (November 2021). "Deciphering mouse uterine receptivity for embryo implantation at single-cell resolution". Cell proliferation. 54 (11): e13128. doi:10.1111/cpr.13128. PMID 34558134.
  7. ^ Teh, WT; McBain, J; Rogers, P (November 2016). "What is the contribution of embryo-endometrial asynchrony to implantation failure?". Journal of assisted reproduction and genetics. 33 (11): 1419–1430. doi:10.1007/s10815-016-0773-6. PMID 27480540.
  8. ^ Sternberg, AK; Buck, VU; Classen-Linke, I; Leube, RE (6 August 2021). "How Mechanical Forces Change the Human Endometrium during the Menstrual Cycle in Preparation for Embryo Implantation". Cells. 10 (8): 2008. doi:10.3390/cells10082008. PMC 8391722. PMID 34440776.
  9. ^ 6.2 Implantation stages from embryology.ch at by the universities of Fribourg, Lausanne and Bern (Switzerland). Retrieved May, 2012
  10. ^ Xiao, Y.; Sun, X.; Yang, X.; Zhang, J.; Xue, Q.; Cai, B.; Zhou, Y. (2010). "Leukemia inhibitory factor is dysregulated in the endometrium and uterine flushing fluid of patients with adenomyosis during implantation window". Fertility and Sterility. 94 (1): 85–89. doi:10.1016/j.fertnstert.2009.03.012. PMID 19361790.
  11. ^ Crha, I; Ventruba, P; Žáková, J; Ješeta, M; Pilka, R; Lousová, E; Papíková, Z (2019). "Uterine microbiome and endometrial receptivity". Ceska gynekologie. 84 (1): 49–54. PMID 31213058.
  12. ^ Moreno, I; Codoñer, FM; Vilella, F (December 2016). "Evidence that the endometrial microbiota has an effect on implantation success or failure". American journal of obstetrics and gynecology. 215 (6): 684–703. doi:10.1016/j.ajog.2016.09.075. PMID 27717732.
  13. ^ Cakmak, H; Taylor, HS (March 2011). "Implantation failure: molecular mechanisms and clinical treatment". Human reproduction update. 17 (2): 242–53. doi:10.1093/humupd/dmq037. PMID 20729534.
  14. ^ a b c d e f g h i j k l m n o p q r s t u v w x y z aa ab ac ad ae Boron, Walter; Emile Boulpaep (2004). Medical Physiology: A Cellular And Molecular Approaoch. Oxford: Elsevier. ISBN 1-4160-2328-3. OCLC 61527528.[page needed]
  15. ^ Ruan, YC; Guo, JH; Liu, X (July 2012). "Activation of the epithelial Na+ channel triggers prostaglandin E₂ release and production required for embryo implantation". Nature medicine. 18 (7): 1112–7. doi:10.1038/nm.2771. PMID 22729284.
  16. ^ Ruan, Y. C., Guo, J. H., Liu, X., Zhang, R., Tsang, L. L., Dong, J. D., Chan, H. C. (2012). Activation of the epithelial Na channel triggers prostaglandin E2 release and production required for embryo implantation. Nature Medicine, 18(7), 1112–1117. doi: 10.1038/nm.2771
  17. ^ a b Margarit, L.; Taylor, A.; Roberts, M. H.; Hopkins, L.; Davies, C.; Brenton, A. G.; Conlan, R. S.; Bunkheila, A.; Joels, L.; White, J. O.; Gonzalez, D. (2010). "MUC1 as a Discriminator between Endometrium from Fertile and Infertile Patients with PCOS and Endometriosis". The Journal of Clinical Endocrinology & Metabolism. 95 (12): 5320–5329. doi:10.1210/jc.2010-0603. ISSN 0021-972X. PMID 20826587.
  18. ^ Carson, D. D.; et al. (2006). "MUC1 is a scaffold for selectin ligands in the human uterus". Front. Biosci. 11 (1): 2903–8. doi:10.2741/2018. PMID 16720361.
  19. ^ Genbacev, O. D.; et al. (2003). "Trophoblast L-selectin-mediated adhesion at the maternal-fetal interface". Science. 299 (5605): 405–8. Bibcode:2003Sci...299..405G. doi:10.1126/science.1079546. PMID 12532021. S2CID 15462671.
  20. ^ Singh, H; Aplin, JD (July 2009). "Adhesion molecules in endometrial epithelium: tissue integrity and embryo implantation". Journal of anatomy. 215 (1): 3–13. doi:10.1111/j.1469-7580.2008.01034.x. PMID 19453302.
  21. ^ Gipson, IK; Blalock, T; Tisdale, A; Spurr-Michaud, S; Allcorn, S; Stavreus-Evers, A; Gemzell, K (January 2008). "MUC16 is lost from the uterodome (pinopode) surface of the receptive human endometrium: in vitro evidence that MUC16 is a barrier to trophoblast adherence". Biology of reproduction. 78 (1): 134–42. doi:10.1095/biolreprod.106.058347. PMID 17942799.
  22. ^ Gardiner, C., Tannetta, D., Simms, C., Harrison, P., Redman, C., Sargent, I., & Milstone, D. (2011). Syncytiotrophoblast Microvesicles Released from Pre-Eclampsia Placentae Exhibit Increased Tissue Factor Activity (Tissue Factor in Pre-Eclampsia). PLoS ONE, 6(10), e26313. https://doi.org/10.1371/journal.pone.0026313
  23. ^ Zhu, JY; Pang, ZJ; Yu, YH (2012). "Regulation of trophoblast invasion: the role of matrix metalloproteinases". Reviews in obstetrics & gynecology. 5 (3–4): e137-43. PMID 23483768.
  24. ^ Carter, AM; Enders, AC; Pijnenborg, R (5 March 2015). "The role of invasive trophoblast in implantation and placentation of primates". Philosophical transactions of the Royal Society of London. Series B, Biological sciences. 370 (1663): 20140070. doi:10.1098/rstb.2014.0070. PMID 25602074.
  25. ^ Moser, G; Weiss, G; Sundl, M; Gauster, M; Siwetz, M; Lang-Olip, I; Huppertz, B (March 2017). "Extravillous trophoblasts invade more than uterine arteries: evidence for the invasion of uterine veins". Histochemistry and cell biology. 147 (3): 353–366. doi:10.1007/s00418-016-1509-5. PMID 27774579.
  26. ^ Dawe, GS; Tan, XW; Xiao, ZC (January 2007). "Cell migration from baby to mother". Cell adhesion & migration. 1 (1): 19–27. PMID 19262088.
  27. ^ Ai Z.; Jing W.; Fang L. (2015). "Cytokine-Like Protein 1 (Cytl1) A Potential Molecular Mediator in Embryo Implantation". PLOS ONE. 11 (1): e0147424. doi:10.1371/journal.pone.0147424. PMC 4723121. PMID 26800213.
  28. ^ Melford, S. E.; Taylor, A. H.; Konje, J. C. (2013). "Of mice and (wo)men: factors influencing successful implantation including endocannabinoids". Human Reproduction Update. 20 (3): 415–428. doi:10.1093/humupd/dmt060. ISSN 1355-4786. PMID 24306146.
  29. ^ a b c Cakmak, H.; Taylor, H. S. (2010). "Implantation failure: Molecular mechanisms and clinical treatment". Human Reproduction Update. 17 (2): 242–253. doi:10.1093/humupd/dmq037. PMC 3039220. PMID 20729534.
  30. ^ Potdar, N.; Gelbaya, T. A.; Konje, J. C.; Nardo, L. G. (2013). "Adjunct low-molecular-weight heparin to improve live birth rate after recurrent implantation failure: A systematic review and meta-analysis". Human Reproduction Update. 19 (6): 674–684. doi:10.1093/humupd/dmt032. PMID 23912476.

Further reading[edit]