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Journal of the Anatomical Society of India

The Effects of Passive Smoking on the Terminal Villi of Human Placenta

Author(s): Rath, G., Jain, A.K*., Bastia, B., Sood M. and Mukherjee A*.

Vol. 50, No. 1 (2001-01 - 2001-06)

Department of Anatomy and Obstetrics & Gynaecology, Lady Hardinge Medical College and S.K. Hospital, New Delhi, *Institute of Pathology, ICMR, New Delhi—INDIA.

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Abstract

One micron thick sections of terminal villi of fifty placentae of passive smoking mothers and equal number of gestationalmatched placentae for control were studied under light microscope. In case of passive smokers, the villi revealed increased syncytial knots,degenerated nuclei in syncytiotrphoblast, hyperplasia in cytotrophoblast, irregular thickening of basement membrane and bulging of theendothelial cells into the lumen of fetal capillaries. The observations were found to be statistically significant in comparison to control andpositively correlated with the findings reported by other authors in the placenta of active smoker. Thus, it may be concluded that passivesmoking by pregnant women also causes the impairment of villous structure and may hamper the materno-fetal transfer leading to deleterious effect on the fetus as noticed in active smokers. Key words : passive smoking, placenta, terminal villi.

Introduction :

Tobacco in any form is injurious to almost all systems of the body (USDHHS Publication, 1998). Its toxic effect on developing fetus, newborn infants and placenta has also been reported (Windham et al, 1992, Demir et al, 1994, Lieberman et al, 1994, Olds et al, 1994). Placenta plays a major role in exchange of toxic substances through circulation. Obvious morphological changes in the placental villi have been noticed depending upon the quantity of tobacco taken by the pregnant women during pregnancy (Asmussen, 1980, Vander Veen and Fox, 1982). It has been reported that lung and brain tumours are also common to passive smokers (Filippini, 2000). The incidence of low birth weight, IUGR, prematurity and stillbirth has been associated with passive smoking (Wenderlein, 1995). Very scanty report is available on the morphological changes in placenta of passive smokers of India. Therefore, the present study is an attempt to find out the histopathological changes in the placenta of the passive smokers under light microscope.

Materials and Methods :

One hundred placentae were collected immediately after delivery (38 - 42 weeks of gestation) from the department of Obstetrics and Gynaecology of Lady Hardinge Medical College, New Delhi. Out of hundred placentae, fifty were collected from women with no history of smoking and other complications during and after pregnancy which were labelled as control and rest fifty placentae from passive smokers. The pregnant mother exposed to tobacco (mostly beedi) used by chain smoking husband or very close relative (30 beedis or more per day) in a nuclear family were designated as passive smoker. Other chronic debilitated diseases like, diabetes, hypertension, tuberculosis have been excluded from our study.

The gestational age and fetal weight were taken from the clinical case-sheets.

Immediately after the expulsion of the placenta after delivery the umbilical cord was cut 5cm away from the site of insertion and membranes were trimmed. The clots were removed from the maternal surface and gently blotted dry with filter paper and finally the weight of the placenta was taken. It was followed by perfusion with 2% glutaraldehyde (pH 7.2-7.4) From maternal surface, two samples (one from center and other from periphery) of about 1 cm thickness were cut from each placenta and fixed in For-glu (pH 7.2-7.4) for 24 hours. Routine processing was carried out for preparation of resin blocks. One-micron thick sections were cut and stained with toluidine blue. As no appreciable change was noticed in the stem villi and intermediate villi, most of our observations were limited to terminal villi. One hundred terminal villi were examined at random in each sample. The histology was studied, significant morphological changes in tobacco exposed placentae were noted down and results were analysed in comparison to control placentae. Due to non-availability of the estimation method, the exact amount of inhalation of tobacco could not be estimated. The study was purely based on the history given by the patient.

Results:

No gross abnormality was noticed in shape, size, calcification or infarction of placental tissue in control or in the passive smoker groups. The placental weight was found to be more in passive smokers than control mothers, which was statistically significant. It was also compared with the corresponding weight of the newborn and was found to be inversely proportional to the weight of the placenta (Table 1). Under light microscope, the toluidine blue stained semithin sections presented a number of terminal villi per field, which were separated from one another by intervillous space containing maternal RBCs. Each of the villi was studied in detail under the following headings and the relevant informations were tabulated and level of significance for all the parameters is also depicted in Table-1.

TABLE-I: COMPARISON OF VARIOUS PARAMETERS OF PLACENTA IN CONTROL AND PASSIVE SMOKER GROUP

Parameters Control
(n=50)
Mean±SD
Passive Smoker
(n=50)
Mean±SD
t-Value
Placental weight (gms) 438 ± 37.12 542 ± 42.36 13.055**
Infant weight (kg) 2.71 ± 0.12 2.33 ± 0.11 13.195**
Histological features (per 100 villi)
syncytial knots (no. of knots) 24.10 ± 5.30 36.50 ± 7.20 9.805**
Syncytiotrophoblast (necrosis and degenerative changes) 10.30 ± 0.52 30.45 ± 6.25 22.715**
Cytotrophoblast (no. of cells) 4.27 ± 1.62 70.25 ± 14.70 31.545**
Endothelial cells (Protrusion into lumen) 5.35 ± 1.47 51.66 ± 12.75 25.515**
Vasculosyncytial membrane (incidence) 26.35 ± 10.70 27.17±9.85 0.4
**: Significant at 5% level of significance.

The syncytiotrophoblast, the outermost lining of the terminal villi, was continuous with each other. The nuclei were irregular in shape and darkly stained with condensed chromatin material inside it. At some places they were scattered in the cytoplasm, and at other places they were present in groups or protrude into the intervillous space forming syncytial knots in both the control and passive smoker groups. It has been noticed that the number of syncytial knots was more in passive smokers (36% approx.) in comparison to control (24% approx). Degenerative and necrotic changes in the nuclei were more obvious in the passive groups (30%) also. The cytoplasm was darkly stained, finely granular and its inclusions were not very clear in semithin sections in both the groups (Fig. 1 and 2).

The cytotrophoblast in case of full-term control placenta was occasionally present under syncytiotrophoblast. But in case of the passive smoking placentae, in almost all villi in a field, the cytotrophoblasts were discernible. The cell outline was very clear, but the nuclei were mostly oval in shape. They were lighter in staining and at some places the division of cytotrophoblast was clearly noticed. This hyperplasia of cytotrophoblast was observed in 70.25% of terminal villi. The cytoplasm appeared granular but detailed inclusions could not be identified in the semithin sections (Fig. 2).

The Trophoblastic Basement Membrane (TMB), in case of the control was thin and was situated between the stroma and the syncytiotrophoblast. Whereas in case of the passive smoking group, it was noticed below cytotrophoblast and was appreciably thicker than control. This thickness was irregularly irregular in about 75% of terminal villi studied. Opposite to the cytotrophoblast, the thickness was found to be more than the area devoid of cytotrophoblast. The TBM in case of passive smoking group was found to be running in a more zig-zag manner than control (Fig. 3).

The Stroma consists of collagen fibres, connective tissues and their nuclei. At some places it was very thick and at other places it was difficult to appreciate between the trophoblastic basement membrane and the fetal capillaries. No appreciable changes were noticed in control and passive smoker group. (Fig 1 and 2).

The Capillaries in both the groups were lined by endothelial cells. In case of control, the lumen of the capillaries appeared smooth and the endothelial cells were running transversely along the margin of the lumen. Whereas in case of passive smoker, the lumen was irregular and endothelial cells were protruding into the lumen of capillaries. In 51% of cases of passive smokers, the protrusion of endothelial cells into the lumen of capillaries was evident. (Fig. 4, 5 and 6).

The Vasculosyncytial membrane (VSM) which is considered to be the principal site for gaseous transfer was the area of the cytoplasm of syncytiotrophoblast in close approximation with the capillary having minimal amount of stroma between the trophoblastic and endothelial basement membrane. In case of passive smoker, the VSM was found to be thicker at the site of cytotrophoblast, whereas in the absence of cytotrophoblast the thickness appeared to be same as control. The incidence of VSM in terminal villi was found to be more or less equal in both the cases. (Fig. 1, 3 and 6).

Discussion :

In India, the smoking habits in low and middle socio-economic class are more common among men than women (W.H.O. report, 1997 and Gupta, 1996). The spouse, child and close associates face the consequence without being directly exposed. As the placenta is the most important organ between mother and fetus during pre-natal period, the toxic effect of nicotine may be manifested in it (Lieberman et al, 1994).

Pfarrer et al (1999) have noticed the increase in placental weight and corresponding decrease in fetal weight in tobacco smoking mother. In our study also it has been observed that the placental weight is inversely proportional to the fetal weight. This may be due to the compensatory mechanism, which involves in the increase in size of the placenta as reported by Sastry (1991) in relation to smoking.

The histopathological changes in placental villi of passive smoking mothers were more pronounced as compared to the non-smokers. In case of controls, the overall structure of the placenta was maintained barring some minor non-significant histopathological appearances, such as syncytial knots, degenerated and necrotic syncytiotrophoblast and few cytotrophoblast.

Demir et al (1994) have noticed increased syncytial knots in the placenta of active smokers as in our study based on passive smokers. Degenerative and necrotic changes in the nucleus of syncytiotrophoblast have been noticed in 30.45 ± 6.25% of villi in the present study. According to Vander Veen and Fox (1982) the necrosis and increased syncytial knots are nothing but the signs of degenerative changes due to ischaemic conditions or hypoxia.

Marana et al (1998) have studied that human syncytiotrophoblast undergoes apoptosis as evidenced by electron microscopic study, but this process is inhibited in case of smoking mother. In our case, it was unable to detect the reduced incidence of syncytiotrophoblastic apoptosis since it was only a light microscopic analysis of the placental villi. But the degenerative and necrotic changes in the syncytiotrophoblast which was noticed in 30.45 ± 6.25% of villi studied in case of passive smokers may be indicative of decreased syncytiotrophoblastic apoptosis in the placenta of smoking mothers as observed by these authors. This inhibiting effect of the smoking habit may be associated with the alterations in the materno-fetal exchange, which could be the possible cause for poor prognosis of the pregnancy in the presence of maternal smoking.

The cytotrophoblastic hyperplasia and degeneration has been noticed by Vander Veen and Fox (1982) in active smokers. In our study, only cytotrophoblastic hyperplasia has been recorded in 70.25 ± 14.70% of villi. This hyperplasia may be due to failure of transformation of cytotrophoblast into syncytiotrophoblast mostly associated with placental hypoxia (Castellucci et al, 1990). In our light microscopic study on placentae of passive smokers, degeneration of cytotrophoblast could not be appreciated.

Marked thickening of the trophoblastic basement membrane is associated with various pathologic conditions, such as pre-eclampsia and essential hypertension (Fox, 1968), maternal diabetes (Liebhart, 1971 and 1974) and IUGR (Macara et al, 1996). In the present study of the passive smoking group, the TBM was also found to be appreciably thicker than the control group. This thickness was a notable feature in about 75% of the terminal villi studied. Thickenng of TBM may be regarded as an important cause contributing to fetal abnormalities and death as it may hamper the transfer of IgG to fetal circulation. According to Benirschke and Kaufmann (1995) the increased thickness of TBM may be due to an increased secretion or decreased turnover of basal lamina molecules.

Burton et al (1989) had observed a significant reduction in the diameter of lumen of the capillaries of terminal villi. In our study, no obvious reduction in diameter was discernible, but the protrusions of endothelial cells into the lumen of capillaries may result in the reduction of the diameter of capillaries in passive smokers. This abnormality was noted in about 51.66 ± 12.75% of the villi studied that might have contributed to the fetal hypoxia and altered fetal haemodynamics. Though Asmussen (1980) has commented on the decrease in number of fetal capillaries in placentae of active smoker group, but in our study group no reduction in number of capillaries was noticed in passive smokers in comparison to control.

Paucity of Vasculo-syncytial membrane (VSM) has been noticed in case of hypertension, preeclampsia and smoking placenta (van der Veen & Fox, 1982, van der Velde et al, 1983, Sodhi et al, 1990, Demir et al, 1994). Whereas in our study, equal incidence of vasculosyncytial membrane in both the groups may be the manifestation of moderate hypoxia resulting due to passive smoking.

Most of our histological findings on terminal villi of placenta of passive smoking mother were similar to the findings observed by other authors on the direct effect of smoking on placenta. Though our study is an indirect correlation, still it highlights the altered placental morphology, which may be hazardous to the developing fetus belonging to passive smokers exposed during antenatal period.

Acknowledgement :

The financial assistance provided by Indian Council of Medial Research (I.C.M.R.), New Delhi to carry out the study is gratefully acknowledged.

References :

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J. Anat. Soc. India 50(1) 19-23 (2001)

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Fig. 1: Terminal villi from a control placenta showing showing darkly stained syncytiotrophoblast (S) and syncytiotrophoblast (S), trophoblastic basement lighter stained cytotrophoblast (C). The trophoblastic membrane (TBM), fetal capillary (F.C.) stroma (St), basement membrane (TBM) separates the stroma (St.) intervillous space (IVS) and vasculosyncytial from the trophoblast. Note the degenerative nucleus membrane (VSM). 500X. (DN) in syncytiotrophoblast, Fetal capillary (F.C.) and intervillous space (IVS). 500X.

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Fig. 2: Terminal villi of the placenta of a passive smoker

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Fig 3: Thickening of the trophoblastic basement membrane (TBM) in passive smoker. Syncytiotrophoblast (S), cytotrophoblast (C), fetal capillary (F.C.), stroma (St), vasculosyncytial membrane (VSM) and intervillous space (IVS). 500X.

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Fig 4: Smooth outline of capillary of the terminal villi from a control placenta, endothelial cell (E), syncytiotrophoblast (S), syncytial knot (S.K.) fetal capillary (F.C.), stroma (St) and intervillous space (IVS). 200X.

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Fig 5: Protrusion of endothelial cells (E) into the lumen of the capillary of a passive smoker. Syncytiotrophoblast (S), cytotrophoblast (C), fetal capillary (F.C.), stroma (st) and intervillous space (IVS). 200X.

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Fig. 6: Difference in thickness of vaculosyncytial membrane (VSM) in placenta of passive smoker. Syncytiotrophoblast (S), cytotrophoblast (C), stroma (St) and intervillous space (IVS). Note the protrusion of endothelial cell (E) into the lumen of the fetal capillary (F.C.). 500X.

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