Molecular Human Reproduction vol.5 no.7 pp. 656–661, 1999
Secretory leukocyte protease inhibitor (SLPI) concentrations in
cervical mucus of women with normal menstrual cycle
Akihiro Moriyama1, Koichiro Shimoya1,2,4, Isao Ogata1, Tadashi Kimura1, Takafumi Nakamura1,
Hiroko Wada1, Kazutomo Ohashi1, Chihiro Azuma1, Fumitaka Saji3, and Yuji Murata1
1Department of Obstetrics and Gynecology, Faculty of Medicine, Osaka University, 2–2 Yamada-oka, Suita City, Osaka
565-0871,2Department of Obstetrics and Gynecology, Osaka Police Hospital, 10–31 Kitayama-cho, Tennouji-ku, Osaka
543-8502, and3Department of Gynecology, Osaka Medical Center for Cancer and Cardiovascular Diseases, 1–3–3
Nakamichi, Higashinari-ku, Osaka 537-0025, Japan.
4To whom correspondence should be addressed at: Department of Obstetrics and Gynecology, Faculty of Medicine,
Osaka University, 2–2 Yamada-oka, Suita City, Osaka 565-0871, Japan. e-mail: email@example.com
Secretory leukocyte protease inhibitor (SLPI) is a potent inhibitor of human leukocyte elastase. SLPI transcripts
in the cervical tissue were detected during the menstrual cycle by reverse transcription–polymerase chain
reaction (RT–PCR). Western blot analysis revealed that the intensity of SLPI protein in cervical tissue in the
ovulatory phase was stronger than in other phases. Immunohistochemistry using an anti-SLPI polyclonal
antibody revealed positive staining in the epithelial cells of the endocervix. Western blot analysis also revealed
that SLPI protein was present in the cervical mucus. Again the intensity of SLPI protein in the ovulatory
phase was stronger than that in the follicular phase. The SLPI concentrations and SLPI/elastase ratios in the
cervical mucus of women in the ovulatory phase were significantly higher than in the follicular and luteal
phases. The SLPI and elastase concentrations in the cervical mucus were positively correlated. No significant
difference was found in the SLPI serum concentrations of women during the menstrual cycle. These results
suggest that production of SLPI from cervical epithelial cells during the ovulatory phase may be important
for protection from the effects of elastase.
Key words: cervical mucus/ovulation/secretory leukocyte protease inhibitor (SLPI)
It is important for the investigation of infertility to evaluate
cervical factor and sperm–cervical mucus interactions. Abnor-
malities of the cervix and its secretion are responsible for
infertility in ~5–10% of infertile women (Moghissi, 1995).
Cervical mucus is a complex secretion produced constantly
by the secretory cells of the endocervix. The cervix produces
mucus at the rate of 20–60 mg/day in normal women of
reproductive age. During the midcycle, the amount increases
10–20-fold and may reach up to 700 mg/day (Moghissi
and Syner, 1976). Cyclic variations in the amount, physical
properties,andchemical contentofthecervical mucusconstitu-
ents have been reported. From the termination of menstruation
to the time of ovulation, viscosity and flow elasticity progress-
ively decrease and spinbarkeit increases. After ovulation and
and viscosity markedly increase (Moghissi, 1995). Cervical
mucus has bacteriostatic and bactericidal properties against
certain strains of bacteria. Various bacteria are unable to
migrate in a capillary tube filled with ovulatory cervical mucus.
Bactericidal activity of the human cervical mucus is present
during all phases of the menstrual cycle but is least pronounced
at ovulation (Enhorning et al., 1970).
Pooled human cervical mucus contains ~1–3% protein in
two basic forms, soluble proteins and mucin (Moghissi, 1995).
The major components of soluble proteins are albumin and
© European Society of Human Reproduction and Embryology
gamma globulin. Cyclic variations in the amounts of several
proteins in cervical mucus have been described. In general,
there appears to be a pre-ovulatory decrease and a post-
ovulatory increase in the amounts of albumin, α1-antitrypsin,
and immunoglobulins (Ig) (Schumacher and Pearl 1968;
Schumacher, 1970). Mucins comprise 45% of proteins in the
cervical mucus. Mucin plays an important role in sperm
transport. The secretion of cervical mucus is regulated by
ovarian hormones. Oestrogen stimulates the production of
copious amounts of watery mucus, whereas progesterone
inhibits the secretory activity of cervical epithelial cells
(Moghissi, 1995). The change of cervical mucus may also
influence sperm penetrability, nutrition, and survival. Pre-
ovulatory mucus is most receptive to sperm penetration
(Moghissi et al., 1972).
Secretory leukocyte protease inhibitor (SLPI) is a potent
inhibitor of human leukocyte elastase, human cathepsin G,
and human trypsin (Thompson and Ohlsson, 1986). The
concentrations of SLPI in biological samples have been mon-
itored tocorrelate theseconcentrations withpathological condi-
tions (Kida et al., 1992; Kouchi et al., 1993; Sluis et al.,
1994). SLPI is found in various fluids, including parotid
secretions (Thompson and Ohlsson, 1986), bronchial, nasal
(Fryksmark et al., 1989), cervical mucus (Casslen et al., 1981;
Helmig et al., 1995), and seminal plasma (Ohlsson et al.,
1995; Moriyama et al., 1998). We have reported the beneficial
SLPI in cervical mucus
effect of SLPI on sperm motility damaged by elastase. No
study of the relationship between SLPI and elastase in cervical
mucus during menstrual cycle has been reported. Evaluations
of SLPI concentrations and elastase titres in the cervical mucus
and SLPI gene transcript in the cervical tissue are necessary.
In this study, SLPI protein was detected in the cervical mucus
and cervical tissue by Western blot analysis. SLPI and elastase
were quantified in cervical mucus from women with normal
menstrual cycles by an enzyme-linked immunosorbent assay
(ELISA). SLPI transcripts were also demonstrated in the
cervical tissue by reverse transcription–polymerase chain reac-
tion (RT–PCR) and SLPI producing cells by an immunohisto-
Materials and methods
A total of 11 non-pregnant women (aged 24–35 years) with normal
ovulatory cycles confirmed by basal body temperature and vaginal
sonography were recruited for this study, and informed consent was
obtained. None of the subjects had a history of venereal infection,
and women infected with bacteria or Chlamydia were excluded.
Ovulation was confirmed by a urinary luteinizing hormone (LH) test,
transvaginal sonography, mid-luteal phase progesterone and basal
body temperature charts. Cervical secretion samples (n ? 155) were
collected with sterile Dacron swabs as previously described (Kanai
et al., 1997); the Dacron swab was used to aspirate 150 µl of cervical
mucus. The samples were diluted with saline for 30 min at room
temperature. Debris and cells were removed by centrifugation at
1000 g for 15 min, and the supernatants were stored at –20°C until
titration to determine the SLPI and elastase concentrations. At the
time of collection of cervical mucus, serum was also collected and
stored at –80°C until the titration of SLPI. For use in a Western blot
analysis, some of the cervical mucus samples in the follicular phase
and the ovulatory phase were obtained by aspiration from the
endocervix with a long tuberculin syringe. It was impossible to collect
cervical mucus in the luteal phase by aspiration with a long tuberculin
syringe. Surgical specimens of the cervix were obtained at a hysterec-
tomy for uterine myoma with informed consent, and fresh frozen
sections were prepared immediately.
The Hela cell lines were purchased from American Type Culture
Collection (Rockville, MD, USA).
Goat anti-SLPI polyclonal antibodies and recombinant SLPI
(Escherichia coli expressed) were purchased from R&D Systems
(Minneapolis, MN, USA). A control goat IgG for the control of a
histochemical analysis was purchased from Zymed Laboratories
(San Francisco, CA, USA).
RNA was extracted from cervical tissue samples of 0.5 g wet
weight and the Hela cell lines by acid guanidine thiocyanate–
phenol–chloroform extraction according to a previously described
method (Chomczynski and Sacchi, 1987). Briefly, cervical tissue
samples were rapidly homogenized in 1 ml of denaturing solution
(4 mol/l guanidinium thiocyanate, 25 mmol/l sodium citrate, 0.5%
sarcosyl, 0.1 mol/l 2-mercaptoethanol) at room temperature. The
viscous solution was then transferred to a 5 ml polypropylene tube.
Sequentially, 100 µl of 2 mol/l of sodium acetate (pH 4.0), 1 ml of
acid phenol (water-saturated), and 0.2 ml of chloroform and isoamyl
alcohol mixture (49:1, vol:vol) were added to the homogenate, with
mixing by inversion after the addition of each reagent. The mixture
was then vigorously shaken and placed on ice for 15 min. Samples
were centrifuged at 10 000 g, the aqueous phase containing the RNA
was transferred to another set of polypropylene tubes and mixed with
1 volume of isopropanol. Precipitation of RNA was performed at –
20°C for 30 min. Total RNA was then sedimented at 10 000 g for
10 min at 4 °C. The RNA pellets were pooled and dissolved in 0.3 ml
of denaturing solution, and the RNA was re-precipitated with 0.3 ml
isopropanol for 30 min at –20°C. The sedimented RNA was washed
in 75% ethanol and air-dried. The dry RNA pellet was resuspended
in 10 µl of diethylprocarbonate water, and the RNA concentration
was assayed by measurement of the optical density at 260 nm.
RT–PCR was carried out using an RT–PCR high kit (Toyobo Co,
Tokyo, Japan). The reaction was carried out in the presence of
and 1 µl RNA sample in a 5? RTase buffer, random primers, and
dNTP mix for 40 min at 42°C. PCR amplification was performed,
using an RT mixture (10 µl), with sequence-specific primers against
human SLPI (5?-ACTCCTGCCTTCACCATGAA-3?/5?-CATTCG-
ATCAACTGGCACTT-3?). PCR was carried out for 35 cycles using
a thermal cycler (Perkin-Elmer/Cetus, Norwalk, CT, USA). Each
cycle consisted of denaturation at 94°C (40 s), annealing at 52°C
(40 s), and extension at 72°C (40 s). The amplification yielded a 570
bp DNA product according to the published sequence of the SLPI
gene (Stetler et al., 1986). PCR products were digested by BamHI to
confirm that they were exact SLPI gene transcripts. After digestion,
the 570 bp DNA product was digested to 336 and 234 bp DNA. RT
was performed with total RNA without reverse transcriptase (a mock
RT sample) to detect possible contamination in RNA samples by
genomic DNA. A 20 µl aliquot of a 50 µl PCR mixture was
electrophoresed on a 4% agarose gel and stained in ethidium bromide,
and amplified products were visualized by UV illumination. Molecular
sizes were estimated using a 100 bp DNA ladder. All primers were
obtained from Becks (Tokyo, Japan).
Tissue preparation for Western blot analysis
The homogenizing buffer for protein extraction from the cervical
tissue consisted of 0.5 M Tris–HCl (pH 6.8), 10% sodium dodecyl
sulphate (SDS), 6% β-mercaptoethanol, and 1% Bromophenol Blue
(BPB). The cervical tissues were homogenized in a 2 ml volume.
Homogenates were centrifuged at 4°C for 30 min at 14 000 g to
remove debris. Following protein determinations, the samples were
divided into aliquots, and subjected to polyacrylamide gel electro-
Protein concentrations were determined with Bio-Rad (Hercules, CA,
USA) Protein Determination Reagent, according to the method of
Bradford (Bradford, 1976).
Western blot analysis of cervical mucus and cervical tissue
To determine SLPI protein in the cervical mucus and the cervical
tissue, Western blot analysis was carried out using an anti-human
SLPI polyclonal antibody. Cervical mucus (10 µl) and ~10 µg
of cervical tissue protein were electrophoresed on a 15% SDS–
polyacrylamide gel and transferred onto nitrocellulose membranes
(0.45 µm; Schleicher and Schuell, Dassel, Germany). The membrane
was incubated with 5% dried milk protein followed by anti-human
A.Moriyama et al.
SLPI polyclonal antibody. The primary antibody was used at a final
using an enhanced chemiluminescence (ECL) Western blotting ana-
lysis system (Amersham, Aylesbury, UK).
Immunohistochemical staining of SLPI in the uterine cervix
To determine the localization of SLPI in the uterine cervix, we
performed immunohistochemical staining using an avidin–biotin per-
oxidase complex method kit (OminiTags Universal Streptavidin/
Biotin Affinity Immunostaining Systems, Lipshaw, Pittsburg, PA,
USA). Fresh frozen sections of the cervix were bleached in 0.3%
hydrogen peroxide to block endogenous peroxidase and covered with
2% goat IgG to minimize non-specific binding. The appropriately
Minneapolis, MN, USA) or control goat IgG for the control was
applied at room temperature and left for 1 h. After rinsing with
phosphate-buffered salinesolution, the sections werefurther incubated
for 30 min with biotin-labelled goat anti-mouse immunoglobulin G,
followed by the addition of avidin–peroxidase complex at 4°C.
Peroxidase activity in the sections was visualized with 0.1%
3,3-diaminobenzidinine-tetrahydrochloride containing 0.02% hydro-
gen peroxide in 0.1 mol/l Tris buffer (pH 7.2). The slides were
counterstained with Mayer’s haematoxylin.
Determination of SLPI in the cervical mucus by ELISA
To determine concentrations of SLPI in the cervical mucus, ELISA
kits utilizing a monoclonal antibody specific for SLPI (R&D Systems,
Minneapolis, MN, USA) were used. The SLPI concentration detection
limit of this kit was 62.5 pg/ml. No cross-reactivity with cytokines,
growth factors, elastase, trypsin, and chymotrypsin could be found
in this kit. The intra-assay variation of the SLPI kit was 4.2–8.0%,
and its inter-assay variation was 4.9–8.0%.
Determination of elastase titre in the cervical mucus by
To measure the titres of elastase in the cervical mucus, ELISA kits
mucus titres of elastase which the kit detected were ?1.0 µg/l. The
intra-assay and inter-assay variation of the elastase kit were 2.7–5.2%
and 4.9–9.5% respectively.
Statistical analyses of SLPI, elastase concentrations, and the SLPI/
test; P ? 0.05 was considered to be statistically significant. The
correlation between SLPI and elastase in cervical mucus was analysed
by simple linear regression.
RT–PCR was performed to determine the expression of the
SLPI gene in the cervical tissue during the menstrual cycle.
Figure 1 shows that SLPI transcripts were present in the
cervical tissue during the menstrual cycle. As shown in
Figure 2, the Western blot analysis detected SLPI protein in
the cervical tissue as a 12 kDa band. The intensity of SLPI in
the cervical tissue in the ovulatory phase was stronger than
that in both the follicular and luteal phases. To identify
the origin of this large amount of SLPI, we performed
immunohistochemical staining of sections of the uterine cervix
in the ovulatory phase, using an anti-SLPI polyclonal antibody.
The cytoplasm of columnar epithelial cells in the endocervical
Figure 1. Reverse transcription–polymerase chain reaction (RT–
PCR) analysis of secretory leukocyte protease inhibitor (SLPI)
mRNA expression in the cervical tissue during the menstrual cycle.
Lane 1, DNA size marker: 100 bp ladder. Lane 2, cDNA from the
cervical tissue in the follicular phase. Lane 3, cDNA (Lane 2)
digested by BamHI. Lane 4, cDNA from mock reverse
transcriptase sample of the cervical tissue in the follicular phase.
Lane 5, cDNA from the cervical tissue in the ovulatory phase.
Lane 6, cDNA (Lane 5) digested by BamHI. Lane 7, cDNA from
mock RT sample of the cervical tissue in the ovulatory phase.
Lane 8, cDNA from the cervical tissue in the luteal phase. Lane 9,
cDNA (Lane 8) digested by BamHI. Lane 10, cDNA from mock
RT sample of the cervical tissue in the luteal phase. Lane 11,
cDNA from the Hela cell lines for the positive control. Lane 12,
cDNA (Lane 11) digested by BamHI.
Figure 2. Western blotting of secretory leukocyte protease inhibitor
(SLPI) protein in the cervical tissue. Cervical tissue protein (~10
µg) was electrophoresed on a 15% sodium dodecyl sulphate
(SDS)–polyacrylamide gel and transferred onto nitrocellulose
membranes. Cervical mucus of women in the ovulatory phase was
electrophoresed on a 15% SDS–polyacrylamide gel. The SLPI
signal was detected as described in the text. Lane 1, positive
control (200 ng recombinant SLPI). Lane 2, tissue lysate from
cervical tissue in the follicular phase. Lane 3, tissue lysate from
cervical tissue in the luteal phase. Lanes 4 and 5, tissue lysates
from cervical tissue in the ovulatory phase.
glands, the subepithelial layers, and cervical mucus were
intensely stained (Figure 3). To examine SLPI protein in the
cervical mucus, we performed a Western Blot analysis. As
shown in Figure 4, SLPI protein was detected as a 12 kDa
band in the cervical mucus. The slightly lower molecular
weight band seen in Figures 2 and 4 is thought to be a
degradation product of SLPI. The intensity of SLPI protein in
the ovulatory phase was stronger than that in the follicular
phase. We determined the SLPI concentrations in the cervical
mucus with a specific ELISA for SLPI. Figure 5 shows the
SLPI concentrations in the cervical mucus of women during
the menstrual cycle. The SLPI titres, elastase concentrations
and SLPI/elastase ratio in the cervical mucus of each phase
during the menstrual cycle were calculated by the average of
those of each phase. The SLPI titres of cervical mucus in the
follicular phase were in the range 7–667 ng/ml (median: 188
ng/ml) and those in the luteal phase in the range 14–520 ng/ml
(median: 110 ng/ml), while those in the ovulatory phase were
SLPI in cervical mucus
Figure 3. Immunohistochemical staining of secretory leukocyte protease inhibitor (SLPI)-producing cells in the cervical tissue. Sections of
the cervical tissue in the ovulatory phase were stained by the avidin–biotin complex method with a goat polyclonal anti-SLPI antibody (A,
C) or control goat immunoglobulin (Ig)G (B, D). The columnar epithelial cells of the endocervical glands were intensely stained. (C) and
(D) represent higher magnification views. A and B, Scale bar ? 100 µm, C and D, scale bar ? 25 µm.
Figure 4. Western blotting of secretory leukocyte protease inhibitor
(SLPI) protein in the cervical mucus. Cervical mucus samples of
women in the follicular phase and the ovulatory phase (each 10 µl)
were electrophoresed on a 15% sodium dodecyl sulphate–
polyacrylamide gel. The SLPI signal was detected as described in
the text. Lane 1, cervical mucus sample of women in the ovulatory
phase. Lane 2, cervical mucus sample of women in the late
follicular phase. Lane 3, cervical mucus sample of women in the
early follicular phase. Lane 4, positive control (200 ng recombinant
in the range 352–1040 ng/ml (median: 880 ng/ml). There is a
significant difference between SLPI concentrations between
the three phases (P ? 0.0001). The elastase concentrations of
cervical mucus in the follicular phase, in the ovulatory phase,
and in the luteal phase were in the range 80–740 ng/ml
(median: 130 ng/ml), from 170–1100 ng/ml (median: 310 ng/ml),
and from 90–690 ng/ml (median: 300 ng/ml) respectively. No
Figure 5. Change in concentration of secretory leukocyte protease
inhibitor (SLPI) of the cervical mucus during normal menstrual
cycles (n ? 11). The mean SLPI titres of cervical mucus in the
follicular phase were in the range 7–667 ng/ml (median: 188 ng/
ml), those in the luteal phase were in the range 14–520 ng/ml
(median: 110 ng/ml) and those in the ovulatory phase were in the
range 352–1040 ng/ml (median: 880 ng/ml). There was a
significant difference between SLPI concentrations in these three
phases (P ? 0.0001).
A.Moriyama et al.
Figure 6. Change in secretory leukocyte protease inhibitor (SLPI)/
elastase ratio in the cervical mucus during normal menstrual cycles
(n ? 11). The mean SLPI/elastase ratio in the cervical mucus in the
ovulatory phase was in the range 0.079–0.462 (median: 0.149),
that in the follicular phase was in the range 0.005–0.279
(median: 0.055) and that in the luteal phase was in the
range 0.006–0.230 (median: 0.036). The difference in the SLPI/
elastase ratios of these three phases was significant (P ? 0.001).
Table I. Concentrations of secretory leukocyte protease inhibitor (SLPI)
during the menstrual cycle
significant difference of elastase titres in the cervical mucus
among in the three phases (P ? 0.09) was observed. As shown
in Figure 6, the SLPI/elastase ratio in the cervical mucus in
the ovulatory phase was in the range from 0.079–0.462
(median: 0.149), while that in the follicular phase was in the
range 0.005–0.279 (median: 0.055) and that in the luteal phase
was in the range 0.006–0.230 (median: 0.036). The difference
in the SLPI/elastase ratios of these three phases was significant
(P ? 0.001). We also examined the SLPI concentrations in
the serum of the subjects. As shown in Table I, there was no
significant difference in the SLPI serum concentrations during
the menstrual cycle. To examine the correlation between SLPI
and elastase concentrations in cervical mucus, a simple linear
analysis was performed. The SLPI and elastase concentrations
in the cervical mucus were positively correlated (y ? 4.4x ?
2220, r ? 0.47, P ? 0.0001).
In the present study, it was found that the SLPI gene is
expressed during the menstrual cycle. No significant difference
in SLPI gene expression was detected during the menstrual
cycle by RT–PCR. However, there was a difference between
the SLPI protein concentrations in the cervical tissue during
the menstrual cycle. Western blot analysis revealed that the
intensity of SLPI in the cervical tissue in the ovulatory phase
was stronger than that in both the follicular and luteal phases.
SLPI gene 5? flanking region has a TATA box and multiple
potential nuclear activator protein-1 (AP-1) binding sites,
which are capable of mediating a specific response to induction
by phorbol esters. However the mechanisms controlling SLPI
gene expression in vivo are unknown (Abe et al., 1991).
An immunohistochemical analysis using anti-SLPI polyclonal
antibody showed that the columnar epithelial cells of the
endocervical glands were intensely stained, indicating that
these epithelial cells were the main source of SLPI in the
cervical mucus. The results presented here of an immunohisto-
et al., 1981).
The cervical mucus contains various kinds of enzymes,
e.g. amylase, alkaline phosphatase, esterase, aminopeptidase,
lactate dehydrogenase, and peroxidase. These enzymes show
a marked pre-ovulatory decrease and post-ovulatory rise in
response to progesterone in the luteal phase. It has been
suggested that assays of some of the enzymes that exhibit a
pre-ovulatory decline and post-ovulatory rise may be used to
predict or detect ovulation (Moghissi, 1995). The change of
SLPI concentrations in the cervical mucus of women during
the menstrual cycle was observed in the present study. The
mean SLPI titres of women in the ovulatory phase were higher
than those of women in both the follicular phase and the luteal
phase. However, there is no difference in the serum SLPI of
the women during the menstrual cycle. These results indicate
that the change in SLPI concentrations in cervical mucus is a
localized reaction in the menstrual cycle. The secretion of
cervical mucus is regulated by ovarian hormones. Oestrogen
stimulates the production of copious amounts of watery mucus,
whereas progesterone inhibits the secretory activity of cervical
epithelial cells. SLPI production by the cervical tissue might
be regulated by ovarian hormones. It was also reported that
antileukoprotease concentrations in luteal phase cervical mucus
were higher than those of the follicular and ovulatory phases
(Casslen et al., 1981), in contrast to the results of this study.
Casslen et al. reported that the latter part of the menstrual
cycle is a period when numerous leukocytes are found in the
uterus, a situation which presumably presents a significant task
for inhibitors like SLPI. The discrepancy might be due to the
different methods used to obtain cervical mucus and the
different methods used to determine the protein concentrations.
Further investigations are necessary to explain the discrepancy
Cervical mucus has bacteriostatic and bacteriocidal proper-
ties. Elastase is a protease which is produced by leukocytes
in the cervix (Moghissi, 1995). Cervical mucus and semen
contain large amounts of elastase (Wolff and Anderson, 1988;
Shimoya et al., 1993). However, no significant difference of
elastase titres in the cervical mucus during the menstrual cycle
was observed here. SLPI is an inhibitor of proteases such as
leukocyte elastase (Ohlsson et al., 1995). In the present study,
the mean SLPI titre and SLPI/elastase ratio of the women in
the ovulatory phase were higher than those of the women in
the follicular or luteal phases. The up-regulation of SLPI plays
SLPI in cervical mucus Download full-text
a defensive role in the epithelial surface of inflammatory lung
diseases (Abbinante et al., 1993). SLPI might protect the
cervical epithelium from the leukocyte protease of cervical
mucus and semen.
The sperm–cervical mucus interaction is an important factor
for fertilization. Cyclic alterations in the concentrations of
cervical mucus may also influence sperm penetrability, nutri-
tion, and survival. Pre-ovulatory mucus is most receptive to
sperm penetration (Moghissi and Syner, 1976). It is usually
inhibited within 1–2 days after ovulation but may persist to a
lesser degree for a longer period (Moghissi, 1995). It has
previously been demonstrated that SLPI recovered sperm
motility reduced by elastase which is contained in the seminal
plasma (Moriyama et al., 1998). In human cervical mucus,
motile spermatozoa have been found 2–8 days after coitus
(Moghissi, 1995). Because the cervical mucus in the ovulatory
phase contains high amounts of SLPI, this molecule might
have an important effect on the sperm penetrability of human
the relationship between SLPI concentrations and cervical
factors in infertility.
This work was supported in part by Grants-in-Aid for Scientific
research (Nos. 20151061, 30203897, 50294062, 70283786, 80301266
and 90093478) from the Ministry of Education, Science, and Culture
of Japan (Tokyo, Japan).
Abbinante, N.J., Simpson, L.G. and Leikauf, G.D. (1993) Neutrophil elastase
increases secretory leukocyte protease inhibitor transcript levels in airway
epithelial cells. Am. J. Physiol., 265, L286–292.
Abe, T., Kobayashi, N., Yoshimura, K. et al. (1991) Expression of the
secretory leukoprotease inhibitor gene in epithelial cells. J. Clin. Invest.,
Bradford, M.M. (1976) A rapid and sensitive method for the quantitation of
microgram quantities of protein utilizing the principle of protein-dye
binding. Anal. Biochem., 72, 248–254.
Casslen, B., Rosengren, M. and Ohlsson, K. (1981) Localization and
quantitation of a low molecular weight proteases inhibitor, antileukoprotease
in the human uterus. Hoppe-Seyler’s Z. Phusiol. Chem., 362, 953–961.
acid guanidium thiocyanate–phenol–chloroform extraction. Ann. Biochem.,
Enhorning, G., Huldt, L. and Melen, B. (1970) Ability of cervical mucus to
act as a barrier against bacteria. Am. J. Obstet. Gynecol., 108, 532–537.
Fryksmark, U., Jannert, M., Ohlsson, K. et al. (1989) Secretory leukocyte
protease inhibitor in normal, allergic and virus induced nasal secretions.
Rhinology, 27, 97–103.
Helmig, R., Uldbjerg, N. and Ohlsson, K. (1995) Secretory leukocyte protease
inhibitor in the cervical mucus and in the fetal membranes. Eur. J. Obstet.
Gynecol. Reprod. Biol., 59, 95–101.
Kanai, T., Fukuda-Miki, M., Shimoya, K. et al. (1997) Increased interleukin-
1 and interleukin-1 receptor antagonist levels in cervical mucus in the
ovulatory phase in comparison with the follicular phase. Gynecol. Obstet.
Invest., 43, 166–170.
Kida, K., Mizuuchi, T., Takeyama, K. et al. (1992) Serum secretory
leukoprotease inhibitor levels to diagnose pneumonia in the elderly. Am.
Rev. Respir. Dis., 146, 1426–1429.
Kouchi, I., Yasuoka, S., Ueda, Y. et al. (1993) Analysis of secretory leukocyte
protease inhibitor (SLPI) in bronchial secretions from patients with
hypersecretory respiratory diseases. Tokushima J. Exp. Med., 40, 95–107.
Moghissi, K.S. (1995) Cervical factor in infertility. In Wallach, E.E. and
Zacur, H.A. (eds), Reproductive Medicine and Surgery. Mosby Book Co,
Chicago, USA, pp. 376–397.
Moghissi, K.S. and Syner, F.N. (1976) Cyclic changes in the amount of sialic
acid of cervical mucus. Int. J. Fertil., 21, 246–250.
Moghissi, K.S., Syner, F.N. and Evans, T.N. (1972) A composite picture of
the menstrual cycle. Am. J. Obstet. Gynecol., 114, 405–418.
Moriyama, A., Shimoya, K., Kawamoto, A. et al. (1998) Secretory leukocyte
protease inhibitor (SLPI) levels in seminal plasma: SLPI restores sperm
motility reduced by elastase. Mol. Hum. Reprod., 4, 946–950.
Ohlsson, K., Bjartell, A. and Lilja, H. (1995) Secretory leukocyte protease
inhibitor in the male genital tract: PSA-induced proteolytic processing in
human semen and tissue localization. J. Androl., 16, 64–74.
Schumacher, G.F. (1970) Biochemistry of cervical mucus. Fertil. Steril., 21,
Schumacher, G.F. and Pearl, M.J. (1968) Alpha1-antitrypsin in cervical mucus.
Fertil. Steril., 19, 91–99.
Shimoya, K, Matsuzaki, N., Tsutsui, T. et al. (1993) Detection of interleukin-
8 (IL-8) in seminal plasma and elevated IL-8 in seminal plasma of infertile
patients with leukospermia. Fertil. Steril., 59, 885–888.
Sluis, K.B., Darlow, B.A., Vissers, M.C. et al. (1994) Proteinase-antiproteinase
balance in tracheal aspirates from neonates. Eur. Respir. J., 7, 251–259.
Stetler, G., Brewer, M.T. and Thompson, R.C. (1986) Isolation and sequence
of a human gene coding a potent inhibitor of leukocyte preteases. Nucleic
Acids Res., 14, 7883–7896.
Thompson, R.C. and Ohlsson, K. (1986) Isolation, properties, and complete
amino acid sequence of human secretory leukocyte protease inhibitor, a
potent inhibitor of leukocyte elastase. Proc. Natl Acad. Sci. USA, 83,
Wolff, H. and Anderson, D.J. (1988) Evaluation of granulocyte elastase as a
seminal plasma marker for leukocytospermia. Fertil. Steril., 50, 129–132.
Received on November 26, 1998; accepted on April 9, 1999