Pertanika 10(2), 159-163 (1987)

Preliminary Observations on Intrauterine Leucocytesof the Pregnant, Non-pregnant and Post-parturient Mice

M. ZAMR1 SAAD1

Department of Veterinary Pathology,University of Liverpool,

P.O. Box 147, Liverpool L69 3BX.

Key words: Leucocytes; uterus; mice.

ABSTRAK

Infiltrasi limposit dan leukosit polimorfonukleus dalam endometrium dan miometrium tikus-tikus yang tidak bunting adalah daripada sederhana hingga tinggi. Infiltrasi yang sama dapat dilihatdalam rahim tikus-tikus yang lepas beranak, tetapi infiltrasi sel makrofaj adalah lebih jelas padatikus-tikus yang tidak bunting. Tiga daripada 6 ekor tikus yang tidak bunting dan 5 ekor daripada 6ekor tikus yang lepas beranak menunjukkan infiltrasi leukosit polimorfonukleus yang rendah dalamsel-sel epitelium. Sel leukosit tidak terdapat dalam rahim dan uri (placentome) tikus-tikus bunting.

ABSTRACT

Moderate to high lymphocyte and polymorphonuclear (PMN) leucocyte infiltrations wereobserved in the endometrium and myometrium of non-pregnant mice. Similar density of leucocyteinfiltrations were observed in post-parturient mice, but there were more macrophages in post-parturient than the non-pregnant mice. Three out of 6 non-pregnant and 5 out of 6 post-parturientmice had low PMN leucocyte infiltration in the luminal epithelium. However, in the placentome andinterplacentomal part of the uterus of the pregnant mice, leucocytes were absent.

INTRODUCTIONIntrauterine leucocyte infiltrations have beenobserved in normal and healthy animals. Theleucocytes collected from the uterus possess thephagocytic activity similar to those leucocytes ofthe peripheral blood (Anderson et al., 1985),and are considered to be part of the non-specificuterine defence mechanism (Frank et aL, 1983).

In the mouse, the leucocytic infiltrationoccurs at a certain stage of the reproductivecycle, particularly during oestrus and seems tobe stimulated by mating (Yanagimachi andChang, 1963). This paper describes and quali-tatively compares the leucocyte infiltration in theuterus of non-pregnant, pregnant and post-parturient mice, to be used as a guide in

diagnosing pathological changes in the uterusduring the various reproductive stages.

MATERIALS AND METHODS

Experimental AnimalEighteen out-bred Swiss albino nvce were

used in this study. They were divided into threegroups, non-pregnant, pregnant and the post-parturient groups with 6 mice in each group.

The 6 non-pregnant mice were selected ran-domly from the stock that had never been bredbefore. The status of their oestrus cycle was notdetermined. Three of the pregnant mice werekilled at 44 gram body weight, while the other 3were killed at 54 gram body weight, appro-

1 Present Address: Faculty of Veterinary Medicine and Animal Science, Universiti Pertanian Malaysia, 43400 Serdang,Selangor, Malaysia.

M. ZAMRI-SAAD

ximately 16 and 19 days of pregnancy respec-tively. The post-parturient mice were killed ateither 21 days (3 mice) or at 28 days (3 mice)after parturition. All mice were killed by thecervical dislocation method.

Experimental ProcedurePost-mortem examination was carried outimmediately after death. Cross-section samplesfrom the right uterine horn (middle portion) ofthe non-pregnant and post-parturient mice werefixed in 10% formalin. Samples were taken fromthe cross-section cuts through a placentome andthe interplacentomal area of the right pregnantuterine horn.

After a fixation period of 24 hours, thesamples were trimmed and embedded in

paraffin wax. Five micron thick sections werestained with haematoxylin and eosin andexamined under light microscopy for the pre-sence of leucocytes in the myometrium, endo-metrium, glandular and luminal epithelium andthe foetal membranes. The presence oflymphocyte like cells and PMN leucocytes werescored as absent (—), occasional (+), low( + + ), moderate (+ + +), high (+ + + +) andprofuse (+ + + + +) according to the methoddescribed by Staples et al (1983).

RESULTSTables 1 and 2 summarise the occurrence ofintrauterine leucocytes in non-pregnant,pregnant and post-parturient mice,Lymphocyte-like cells were identified under light

TABLE 1The occurrence of lymphocytes and macrophages in the uterine wall of the non-pregnant,

pregnant and post-parturient mice

Mousenumber

Reproductionstatus

Epithelium Endometrium Myometriumluminal glandular

1.

2.

3.

4.

5.

6.

7.

8.

9.

10.

11.

12.

13.

14.

15.

16.

17.

18.

Not pregnant

Not pregnant

Not pregnant

Not pregnant

Not pregnant

Not pregnant

Pregnant (44g)

Pregnant (44g)

Pregnant (44g)

Pregnant (54g)

Pregnant (54g)

Pregnant (54g)

Post-par (21d)

Post-par (21d)

Post-par (21d)

Post-par (28d)

Post-par (28d)

Post-par (28d)

+ + +

+ +

+ + +

-I- +

•¥• + +

Post-par = post-parturientg ~ gram bodyweightd ~ days after parturitiont = placentome and interplacentome

160 PERTANIKA VOL. 10 NO. 2, 1987

Mousenumber

PRELIMINARY OBSERVATIONS ON INTRAUTERINE LEUCOCYTES OF MICE

TABLE 2The occurrence of PMN leucocytes in the uterine wall of the non-pregnant,

pregnant and post-parturient mice

Reproductionstatus

Epitheliumluminal glandular

Endometrium Myometrium

1.

2.

3.

4.

5.

6.

7.

8.

9.

10.

11.

12.

13.

14.

15.

16.

17.

18.

Not pregnant

Not pregnant

Not pregnant

Not pregnant

Not pregnant

Not pregnant

Pregnant (44g)

Pregnant (44g)

Pregnant(44g)

Pregnant (54g)

Pregnant (54g)

Pregnant (54g)

Post-par (21d)

Post-par (21d)

Post-par (21d)

Post-par (28d)

Post-par (28d)

Post-par (28d)

Post-par = post-parturientg = gram bodyweightd = days after parturitiont == placentome and interplacentome

microscopy as small, rounded cells with a uni-formly stained nucleus and little cytoplasm.Both lymphocytes and macrophages are classi-fied under lymphocyte-like cells. Polymorpho-nuclear (PMN) leucocytes were readily identifiedby their lobulated nuclei and eosinophilic cyto-plasm (Fig. 1).

Non-Pregnant MiceLymphocytes were present at moderate densityin the endometrium and myometrium, parti-cularly in the longitudinal muscular layer ofmost of the non-pregnant mice. They appearedin the uterine wall intermixed with PMN leuco-

Fig. 1: Endometrium of a non-pregnant mouseshowing moderate density of lymphocytes(large arrows) and low density of PMNleucocytes (small arrows). H & E x 400.

PERTANIKA VOL. 10 NO. 2, 1987 161

M. ZAMRI SAAD

cytes, which are present at moderate to highdensity in the endometrium and myometrium.Three out of 6 non-pregnant mice hadoccasional to low PMN leucocytes in the luminaland glandular epithelium (Table 2).

Pregnant MiceNone of the 6 pregnant mice, either killed at 44gram or 54 gram body weight, had any leucocytein the placentome or interplacentomal area(Tables 1 and 2). However, there was a lowincidence of lymphocytes and PMN leucocytes inthe distended and congested placentomal bloodvessels of most of the pregnant mice.

Post Parturient MiceAll three mice killed 21 days after parturitionhad occasional to low PMN leucocytes in theluminal epithelial cells. The endometrium, how-ever, contained occasional PMN leucocytes withmoderate to high density of lymphocyte-likecells. The myometrium contained low tomoderate density of PMN leucocytes, lympho-cytes and macrophages.

Only 1 of the 3 post-parturient mice killed28 days after parturition had low number ofPMN leucocytes in the luminal epithelial cells.All 3 post-parturient mice killed 28 days afterparturition had a high number of lymphocytesand macrophages and low PMN leucocyte intheir endometrium and myometrium.

DISCUSSIONPrevious studies have shown that leucocyte infil-tration in normal uterus occurs at a certain stageof the reproductive cycle. In non-pregnantmouse and rat, it occurs toward the end of theoestrus period (Allen, 1931), and seems to bestimulated by mating (Yanagimachi and Chang,1963). In this study, moderate to high density ofleucocyte infiltrations were observed in theuterus of most of the non-pregnant mice. Thepresence of leucocytes in normal non-pregnantuterus is important as a nonspecific uterinedefence against microbial invasions, particularlyat oestrus, during which stage the female repro-ductive tract is 'open' to many foreign particles(Franks al (1983).

During pregnancy, progesterone is essentialfor the maintenance of pregnancy. It reduces

myometrial contraction, inhibits granuloma re-actions against foreign body implant andreduces the ability to protect against intrauterinebacterial infections (Staples et ai, 1983). Thelatter effect is thought to result from proges-terone-dependent reduction of uterine leucocyteinfiltration (Broome et al., 1960), which wasobserved in the pregnant mice in this study.Acute deprivation of progesterone during earlypregnancy, however, results in a rapid influx ofPMN leucocytes into endometrium, leading tothe degradation of the foetal syncytium (Staplesetai, 1983).

Leucocyte infiltration of the uterine wall inthe post-parturient animals is important for theremoval of post-parturient placental debris(Frank et al., 1983) and bacterial contamina-tions (Eduvie et al., 1984). In this study, most ofthe leucocytes that were classified under'lymphocyte-like cells' in the post-parturientmice were macrophages. Frank et al. (1983) haveobserved the important role played by macro-phages in the involuting uterus. Eighty percentof the phagocytosis of foreign material innormal, involuting uterus is due to PMN leuco-cytes while 20% is due to macrophages.

ACKNOWLEDGEMENTSThe author would like to thank Prof. D.F. Kellyof the Department of Veterinary Pathology,University of Liverpool, for his guidancethroughout the study. Financial support fromthe Association of Commonwealth Universitiesand Universiti Pertanian Malaysia is gratefullyacknowledged.

REFERENCES

ANDERSON, K.L., N.A. HEMEIDA, A. FRANK, H.L.WHITMORE and B.K. GUSTAFSSON. (1985): Col-lection and phagocytic evaluation of uterineneutrophilic leucocytes. Theriogenology 24:305-317.

BROOME, A.W., AJ. WINTER, S.H. MCNUTT andL.E. CASODA. (1960): Variations in uterine res-ponse to experimental infection due to hormonalstate of the ovaries. II. The mobilization ofleucocytes and their importance in uterine bacte-ricidal activity. Am.]. Vet. Res. 21: 675 - 683.

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PRELIMINARY OBSERVATIONS ON INTRAUTERINE LEUCOCYTES OF MICE

EDUVIE, L.O., D.I.K. OSORI, P.B. ADDO, and C O . uterus after acute removal of ovarian progeste-NjOKU. (1984): Bacteriological investigation of rone during early pregnancy in the sheep.the post-partum uterus: Relationship to involu- Placenta 4: 339- 350.

tion and histopathological findings. Theriogeno- YANAGIMACHI. R. and M.C. CHANG. (1963): Infiltra-

£9 tion of leucocytes into the uterine lumen of theFRANK, T., K.L. ANDERSON, A.R. SMITH, H.L. WHIT golden hamster during the oestrus cycle and

MORE and B.K. GUSTAFSSON. (1983): Phagocyto- following mating./ Repr. Fert. 5: 389-396.sis in the uterus: A review. Theriogenology 20:103-110.

STAPLES, L.D., R.B. HEAP, F.B.P. WOODING and G.J.

KING. (1983): Migration of leucocytes into the (Received 28June, 1986)

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