6.2.  The  secretory immune system in human fetuses                          

 

6.2.1)   Lymphoid-epithelial components of the secretory immune system  in  self-protection of human fetuses 

 

Some researchers are of the opinion that the SIS evolves after birth as a reaction to massive microbial invasion and the introduction of large amounts of different foreign antigens through the mucous membranes of the upper respiratory, digestive and urogenital tracts  (60,61). However, as has been shown by the other authors and stated in previous chapters of this book, components of the SIS, such as SC, J chain, IgM, IgA and  lymphocytes, are already present not only in human fetuses in the third trimester of gestation (62,63), but also in embryos of the first trimester (64,65).

 

Maturation of the immune system is well known to start  early in human fetal life (65).  As noted at the beginning of this chapter, B lymphocytes develop in the liver by 9 weeks of gestation and are present in the blood and spleen by 12 weeks. From 14 weeks, T lymphocytes leave the thymus, and subsequently cells with helper and suppressor phenotypes appear in the spleen. The lack of secondary lymphoid tissues in healthy fetuses most probably reflects a lack of antigen stimulus. On the other hand, newborn plasma contains adult levels of IgG which are acquired across the placenta from the mother.

 

In human fetuses,   components of the SIS have been found in the epithelium of the salivary glands and mouth (66,67), trachea and lungs (68,69), and digestive tract (70,71). Small amounts of SC appear in the intestinal mucosa before week 29 of gestation, and  its quantity increases rapidly thereafter (72). Secretory IgA-containing epithelial cells have been found in the respiratory tract and intrahepatic bile ducts of fetuses at 20 to 21 weeks of gestation (73). Immunocompetent mucosa-associated cells (dendritic cells, T lymphocytes, B lymphocytes, and macrophages) have been  found in the human fetal larynx after  week 14 of gestation (74).  SC of   fetal urogenital origin has been found in the amniotic fluid (75). Lymphoid cells expressing   IgA and IgM, as well as other  immunocompetent cells, have been described in fetuses of the second trimester of gestation (76), particularly, in the gut and amniotic fluid (77). From 11 to 14 weeks of gestation, CD68⁺ and CD40⁺ cells   are present throughout the lamina propria. With the emergence of lymphoid aggregates (14-16 weeks), dendritic cells  and B lymphocytes are detected in the fetal gut; however, their expression is restricted to the lymphoid aggregates. Lymphoid follicles forming after 16 weeks of gestation contain  MHC II-positive cells of different subtypes of T lymphocytes.

 

Components of the SIS during the second trimester of gestation (weeks 13-25)  were studied  in 36 human fetuses obtained as a result of medically or socially recommended abortions (group I) or which had died of different causes -- abruptio placentae, placenta previa, and chorioamnionitis (group II) (78). The first group included 21 cases with no signs of foreign  antigenic influences. The second group included 15 cases of acute chorioamnionitis with sepsis, aspiration syndrome or meningitis. In both groups, fetuses were of similar  gestational ages.

 

 

Fetuses without antigenic effects (group I)

 

Elements of the SIS were widespread in the different   organs of these fetuses. SC, J chain, IgA, IgM and IgG were found in the  epithelium and glands of the digestive organs such as the mouth cavity, pharynx, esophagus, stomach and intestine,  throughout the respiratory tract (larynx, trachea, lungs) and urinary tracts (kidneys, ureters, bladder), in hepatocytes and the epithelium of the bile duct, and in acini and ducts of the pancreas, in the follicular epithelium of the thyroid and the ovaries, in the epithelium of the Fallopian tubes and uterus, in the epididymis and rete testes, in the epithelium of the choroid plexuses of the cerebral ventricles, in the mesothelium of the pleura, epicardium and peritoneum, and in the epithelium of the skin, sweat and sebaceous glands. A few macrophages, B cells and different subsets of T lymphocytes   were observed in these structures.  All of these SIS elements were already observed in 13-week-old fetuses and were maintained during the whole second trimester of gestation.

 

The immunoreactivity of the different SIS components varied in different organs. The SC and J chain usually reacted intensively, except for cells in the distal part of the hypophysis and in pancreatic  islands, where the SC was weakly reactive. In Leydig's cells of the testes  and some other organs, where only the J chain was found, the SC was absent. Expression of IgA, IgG and especially of IgM, was very low in the large intestine and hepatocytes in the center of liver lobules. IgM was sometimes  absent in the choroid plexuses of the brain. SIS components   were not seen in the gray substance of the brain, myocardium, intestinal goblet cells, skeletal muscles, fibroblasts, chondrocytes or osteoblasts. A small number of B lymphocytes expressing IgA and IgM was seen in different organs (Table X). 

 

 

 

 

Fetuses with antigenic effects (group II)

 

In  fetuses which had been subjected to massive  antigenic effects at chorioamnionitis, expression of the SC and J chain  was no different from that observed in group I (Fig. 7). Immunoreactivity of IgA, IgG and especially  IgM   was weak or even absent in the epithelium of the skin, respiratory, digestive and urinary tracts, hepatocytes, tubules of the kidneys, and choroid plexuses of the brain. The number of IgA- and IgM-positive lymphocytes increased in the spleen and lymph nodes, lungs, and in the mucous membranes of the stomach and intestine (Table X).

 

It has been shown that the whole SIS-protein-complex is already present  in many tissues of the human fetus  in the second trimester of gestation (78). Such tissues include mainly the widely spread   border tissues covered with the epithelium or its analog, such as the mesothelium of serous cavities.    Our findings of the SIS protein components in the fetal respiratory, digestive and urogenital tracts are very similar to those observed in children and adults (63,79).

 

In fetuses, there are cellular components of the SIS, such as lymphocytes of different subsets, including IgA- and IgM-containing B cells,  and macrophages. However, their amounts are small (Table XI),  and they are diffusely located in subepithelial tissues and intraepithelial spaces. Special lymphoid-epithelial structures, such as tonsils, solitary follicles (nodules) and aggregated follicles (Peyer's patches), which are typical of  the adult SIS (62), were not found in second-trimester fetuses. Such peculiarities in the cellular structure of the fetal SIS as   part of the common lymphoid system of fetuses can be explained by its "immaturity".

 

The total mass of lymphoid tissue in second-trimester fetuses is minute. The total weight of the main lymphoid organs such as the spleen and thymus  is 0.092±0.02% of  mean  body weight in 13- to 15-week-old fetuses, 0.36±0.07% in 23- to 25-week-old fetuses, and 0.73±0.09% in 38- to 40-week-old fetuses. Lymph nodules are few and are of microscopic size. Some lymphoid tissue structures are absent. Reactive centers of lymph nodules are not seen, even in the infected fetuses. The process of transformation of Ig-synthesized B lymphocyte stops at the immunoblast  stage,  and these never become mature plasma cells in cases with acute infections (80).

 

The massive antigenic effects under chorioamnionitis (group II fetuses) caused a two- to fourfold increase in the number of IgA- and IgM-synthesizing lymphocytes in the spleen, lymph nodes, and respiratory and digestive tract organs  where SIS is localized and where the antigenic effect is manifested at its highest level (Table X). As a result, Ig synthesis was activated in the common immune system and in the SIS. The immunoreactivity of IgA, IgM and IgG in the epithelial cells sharply decreased or disappeared altogether. Sometimes, IgA and IgG immunoreactivity   was seen in the fibrin in the bronchial cavity or in the stomach mucus   as a manifestation of the exocrine secretion of Igs  by the epithelium. The described changes could be considered a morphological manifestation of SIS functional activity in human fetuses in the second trimester of gestation.

 

The components of the fetal SIS,  such as IgA and  IgM, are of  fetal origin (60). Maternal IgG has been found in the fetal epithelium, and this is considered  an additional proof  of insufficient functional activity of both the common and secretory fetal immune systems (78). Under massive antigenic attack, immunoreactivity of Igs in the fetal epithelial cells decreases to  complete disappearance, whereas reactivity of the SC and J chain does not change  (Fig. 7). It can be supposed that SC and J chain do not leave the cells during SIS function, and their discovery in the cavities of some organs (such as the amnion,  75) may result from cell shedding and destruction. Cells without exocrine secretion in some organs, such as the distal lobe of the hypophysis, pancreatic islets, etc., contain only the J chain without SC. Our study demonstrated that the  SIS  is widely present and functionally active in   fetuses in the second trimester of gestation. It is not restricted to mucosal membranes, it is present  in the fetal organs  (Fig. 8)  and it plays an important role in the immune defense of the entire fetuses and its strategically important organs against foreign antigenic influences. 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

(Color Figs.)

 

Fig. 7.

 

A.  A 22-week-old fetus. SC in the epithelium of sebaceous glands of the skin (brown staining). ´200.

B. A 19-week-old fetus. J chain in the  epidermis of the skin and the epithelium of sebaceous glands. ´200.

C. A 23-old fetus. Chorioamnionitis. SC in the epithelium of the bronchus (b), esophagus (e), thymus and thyroid (t).  ´100.

D. A 17-week-old fetus. Chorioamnionitis and aspiration syndrome. SC in the epithelial cells of the bronchiole. ´200.  

 

 

 

 

 

 

 

 

 

Fig. 8.

    

A.  A 17-week-old fetus. The kidneys.  SC in the epithelium covering of glomerulus (heads of arrows),  loop of Henle (small arrows), collecting tubules (ct). x100.  

B. The same case.   IgA   in the epithelium of convoluted (small arrows) and collecting (a large arrow) tubules. x200. 

C. A 20-week-old fetus.  The bladder.   SC   in the superficial epithelium. x400.  

D. A 21-week-old fetus. The uterus.  J chain   in the epithelium. x400.   

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Table X.

The number of IgA- and IgM-positive lymphocytes in different fetal organs

(per 50,000 µm², mean ± SE) (After ref. 65)

 

 

^a Significantly different from Group I,  p < 0.05.

Groups of fetuses: I, without antigenic attacks; II, with antigenic attacks.

 

 

 

 

 

 

Table XI. The number of immunocompetent cells in the liver of embryos and fetuses  (mean±SE in 50,000 μm²) (After ref.  65)

 

 

^a Groups of patients: see footnotes to X.

 

 

 

  6.2.2) Components of the secretory immune system in fetal membranes

                                                     and decidua

 

Researches performed to date have been restricted to the study  of single   components of the SIS in human fetal membranes or the placenta. We are not aware of any report on the role of SIS in the whole complex of the maternal-fetal barrier in the first trimester of pregnancy. Because of its ontogenic importance , we investigated the distribution of SIS components in fetal membranes (amnion, yolk sac, chorion) and the decidua throughout the first trimester of pregnancy (64). Specimens from 17 embryos (3.5-4 to 8 weeks of pregnancy) and 9 fetuses (9 to 12 weeks) were divided into those that had not been exposed to massive foreign antigenic effects (group I) and those that had suffered acute chorioamnionitis (group II).

 

Embryos and early fetuses without massive antigenic effects (group I)

 

The presence of SIS protein components (SC, J chain and Igs)  was found in tissues of the gestational sac of both fetal and maternal origin  as early as the fourth  to fifth week of development, and throughout  the rest of the first trimester of pregnancy. In 3.5-4- to 6-week-old embryos not exposed to massive foreign antigenic influences,   SC, J chain and IgG were observed in the epithelium of the amnion, in the syncytio- and cytotrophoblast of the chorion and in the yolk sac endoderm (Fig. 9). CD68+ macrophages were found to contain  J chain, IgA and IgG (Tables XII, XIII), but not SC or IgM. Lymphocytes were not found. Decidual cells   were reactive to SC, J chain and IgA. Single lymphocytes and macrophages were seen in the stroma of the decidua, and only a few of them were positive to IgG, IgA and IgM.

 

In 7- and 8-week-old embryos, immunoreactivity of SC, J chain and IgG was not changed in the amniotic epithelium, endoderm of the yolk sac, trophoblastic and decidual cells. A slight increase in the reactivity of IgA was seen in the latter cells. Groups of CD3+ T lymphocytes and CD20+ B lymphocytes  were seen in angioblasts of the yolk sac  and the aorta  at week 8, while the other subsets of T cells and Igs-containing cells were not found (Table XII).

 

In fetuses at 9 to 12 weeks of pregnancy, immunoreactivity of SC, J chain and IgG was not changed in the above-mentioned tissues, whereas reactivity to IgA and IgM increased. The number of macrophages increased in the stroma of trophoblastic villi (Table XIII). At week 9, single T and B lymphocytes were observed, and at weeks 10 to 11, IgA+ and IgM+ lymphocytes were found. The number of different subsets of lymphocytes increased in the decidual tissue (Table XII). Fibrotic avascular villi were seen in a case  with acardius amorphus. SC, J chain, IgA and IgG were found in the amnionic epithelium and in the trophoblastic and decidual cells.

In fetuses during the last trimester of pregnancy, the amniotic fluid contains, in addition to the predominant maternal IgG, different molecular forms of fetal Igs (81). IgG is found to be the major isotype and to contain mother-derived tetanus antitoxins. IgA is much less abundant, whereas no IgM can be detected. Amniotic fluid of human fetuses at a gestational age of 26 to 40 weeks contains  low levels of IgA of fetal origin and SC in its free form (75,82).   IgG, IgA, and SC are detected in the fetal urine and, therefore, can reach the amniotic fluid via this route. Their function as an immune barrier against infection and against mother-derived autoantibodies has been suggested.

 

Embryos and early fetuses with massive antigenic effects (group II)

 

Massive antigenic exposure due to chorioamnionitis   during the first trimester  caused little change in the distribution and immunoreactivity of SC and J  chain in the amniotic epithelium, trophoblastic and decidual cells. SIS protein components were even found in embryos with heavy abnormalities such as acardius amorphus. Reactivity to IgG and IgA decreased to its disappearance, while reactivity to Igs increased significantly in the perivillous fibrin. Lymphocytes were seen only after week 9 whereas macrophages were observed in high numbers as early as the fourth week of pregnancy  (Table XIII). In the decidual tissue, the number of all types of immunocompetent cells, including Igs-synthesizing lymphocytes and plasma cells, increased sharply (Table XII).

 

The cellular components of the SIS in all fetal membranes  of 3.5-4- to 8-week-old embryos  were represented only by macrophages. CD3+ and CD20+ lymphocytes were seen to the end of week 8, and  IgA+ and IgM+ lymphocytes were found only at weeks 10 to 11. This is in accordance with other authors' observations regarding the development of B cells and Ig-synthesizing lymphocytes (18).

 

The Igs seen in the embryonic tissue appeared to be of maternal rather than embryonic origin. The placental barrier has been shown to be permeable  for maternal IgG (17,19,83).  Specific IgG and IgA were found in the coelomic fluid of 6- to 12-week-old embryos and fetuses suffering from rubella, cytomegalovirus and Toxoplasma gondii infections in amounts similar to their concentrations in the maternal blood (16).  These results suggested that maternal IgG and IgA are potentially available to the embryo as early as week 6 into development. This is in accordance with our  findings that IgG and IgA but not IgM are present in macrophages of the chorionic villi.

 

Macrophages from different   embryonic and fetal organs of the first trimester of pregnancy  were found  to be immunoreactive to IgG and IgA but not to IgM (65). We therefore suggested the presence of transport and defensive functions among macrophages. Thus, during the embryonic period, when the common (systemic)  immune system is still only beginning to develop, Igs of maternal origin appear to be functioning  in the embryonic SIS.

In the decidua, cellular components of the SIS exhibit a full  immune  response during the entire first trimester of pregnancy (84). The small amount of each type of these cells (Table XII) suggests that this response is manifested very weakly,   reflecting mainly  the reaction of a pregnant mother (her decidua) to embryonic antigens.  In the decidua, the number of lymphocytes, including those synthesizing IgG, IgA and IgM, plasma cells and macrophages increases significantly, even during the earliest embryonic period. Such an increase was found even relative to their amount in adult ruptured uterine tubes   (Table XII).

 

Chorioamnionitis with accompanying  massive antigenic effect is characterized by changes in protein and cellular components of the SIS. Igs disappear from the fetal membranes and decidua. With chorioamnionitis, the cellular composition is different in  the embryonic and maternal parts of the gestational sac. In fetal membranes, only macrophages were found to react to the antigenic effect. The absence of a response by lymphocytes of the studied subsets showed that in 3.5-4- to 8-week-old embryos, even a massive antigenic effect  does not accelerate  lymphocyte maturation. A weak lymphocytes reaction   was seen after only 9 to 10 weeks of pregnancy. In the maternal decidua with chorioamnionitis, immune-response development was analogous to an intensive immune reaction in adults (84).

 

Data from the literature (85) and our observations (64,65) show a different origin  and composition of immunocompetent cells and a different course of immune reactions in embryonic and maternal parts of the gestational sac. We conclude  that two   SIS are present at the border between the maternal and embryonic tissues. These systems are already in place at the beginning of the embryonic period, weeks 4 to 5, function during the entire first trimester of pregnancy and are the main immune mechanism underlying the barrier between these two organisms. We suggest that the SIS   plays an important role in the regulation of pregnancy and in the  development of fetal tolerance.

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

(Color Fig.)

 

Fig. 9. 

A 4-week-old normal human embryo. Microphotographs of the gestational sac.

A. SC  (brown staining) in the  decidual cells (d) and invasive trophoblast   (it).  ´100. 

B.  J chain in both the cytotrophoblast and syncytiotrophoblast   and in the macrophages (heads of arrows). x200.

C.  IgG in the syncytiotrophoblast and macrophages (heads of arrows). x200.

D.  IgA in the syncytiotrophoblast and cytotrophoblast and in the macrophages (heads of arrows). x400. 

E.                  Gestational sac of a 8-week-old embryo. SC in the amniotic epithelium (a) and yolk sac endoderm (heads of arrows). A blood island (a white arrow). x200.