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Protocol for the examination of placentas of monozygotic twins, to include those of the TTTS
K. Benirschke June 16, 2000


It is difficult to examine the placenta of the transfusion syndrome for many reasons. In the first place, it is often disrupted and this makes injection study difficult. Moreover, these placentas often are from premature deliveries with placentas that are normally more delicate. Most important is that the examiner is familiar with the distribution of blood vessels in the normal placenta so that he/she can assess the sometimes very hard-to-see anastomoses that are the basis for the transfusion syndrome. Thus he/she should become familiarized by examining normal singleton placentas beforehand.

The transfusion syndrome happens essentially ONLY in monochorionic twin placentas. They may be monoamnionic (MoMo) or diamnionic (DiMo). One of the first requirements is that this status is assured. Thus, section of the “dividing membranes: of DiMo placentas must be made for the record.


PROCEDURE:
At delivery, the umbilical cords of twins (A,B) need to be labeled with one or two clamps and this needs to be recorded and compared with their birth weights. Next, the lengths of the cords are measured (all of it) and recorded. Next, the insertion site of the cords must be documented (central, eccentric, marginal, velamentous). The number of blood vessels in e cord needs to be recorded, then the cords should be detached from the placental mass. The “dividing membranes”, if present, needs to be sampled for histology. The peripheral membranes should then be trimmed and then the placenta is weighed and measured. Then the maternal surface should be inspected for 1) completeness and/or disruption (it helps to know for the later purpose of injection), and 2? Color. It is especially relevant to note if both fetuses’ halves have the same darkness/lightness because this reflects the amount of fetal hemoglobin (thus, this is an indication of anemia and/or plethora).

Next is the fetal surface, the most critical part. The examiner should assess roughly what portion of placental mass is perfused by each twin. Is it 50:50 or is it 80:20, etc. This can be estimated by identifying the “placental vascular equator”. When one pursues, visually, the distribution of blood vessels as they emanate from the insertion of the cords to the periphery the “equator” becomes obvious. At their ends, the fetal blood vessels dip into the villous tissue where they then perfuse a fetal cotyledon. Arteries normally cross over the veins and are thus easily identified (especially in their large calibers near the cords). In the normal placenta, an artery leads to such a peripheral cotyledon, dips down, perfuses the cotyledon, and a corresponding vein brings the blood back from this cotyledon to the same fetus. One needs to identify BOTH these vessels and assure oneself that they come and go to/from the same umbilical cord.

The commonest communication between two monochorionic twins is a single artery-to-artery (A-A) connection on the surface of the placenta; these do NOT dip into villous tissue but are major shunts between the twins and blood can shift back and forth from one twin to the other, depending on blood pressure, heart rate, etc. A vein-to-vein direct anastomosis is uncommon. The A-A anastomosis is of variable size, often it is quite large. But these communications may also be so small that they are hard to detect. It often helps to push blood gently back and forth in the surface vessels and thus make visible small anastomoses, but at times it requires injection.

THE MOST
important communication that is the major responsible agent for the development of TTTS is one where at lest one cotyledon is shared. That is to say, an artery comes from one twin (the donor), dips into a cotyledon, which is then drained by a vein into the other fetus (the recipient), who gets to be larger and plethoric. VERY careful inspection of the fetal surface can actually identify these “common villous districts” (the shared cotyledons). At times though it requires injection. What’s worse, such shared cotyledons may be multiple, of different size, and they may have different blood flow directions. Thus, I always draw a picture of the surface blood vessels and/or photograph them. When I am uncertain or want to demonstrate the communications, then I inject them with milk.


INJECTION
It is foolhardy to try to inject the entire placenta as it is most commonly disrupted somewhere and this defect allows fluid to drain and thus prevents cotyledonary filling. I use milk because it is readily available, can be easily seen when the vessels and villi are filled, and it does not destroy the tissue for later histologic study. I use a completely filled 20 ml syringe with an 18 gauge needle. Then I identify the potential major A-A region or shared cotyledon, so identified from inspection alone, choose it and try to inject this cotyledon only. The nearest feeding (or draining) vessels of adequate size is chosen for injection. One must be careful to insert the needle almost horizontally to the placental surface so as to prevent it from being double punctured. I shove the needle in the chosen vessel for some distance, occlude the vessel around the needle with my fingers and then slowly inject as much milk as is needed to fill the entire cotyledon, until the blood/milk mixture emanates from the other vessel that leads to this shared cotyledon. (If later one inspects that maternal surface or, later still, sections this region, one finds this one cotyledon to be white, as all the blood has been replaced by milk). I record the findings carefully, best by drawings. It is these vessels (actually only one of them) that needs laser occlusion in the TTTS therapy.

A problem arises when one twin has died prior to the twin delivery. When the death has been recent, the anastomoses may still be visible or even injectable. But, when fetal demise has occurred a long time prior to delivery, then injection is fruitless and only very diligent inspection may identify the possible offending blood vessel connections. This may be true even when it is a large blood vessel through which one twin may have bled into the dead twin; they can be most difficult to identify.


MICROSCOPY
Routine sections can be made, but they rarely add to unraveling the syndrome. True, the donor twin’s villi usually are much smaller and those of the recipient are larger and congested; rarely on may find a thrombus, but these findings are of minor consequence; Perhaps the most useful histology comes from a study of the amnionic surface, as the “stuck” twin (donor) often has amnion nodosum, not present in the recipient.

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