Methods
This was a pseudo-longitudinal study (a variant of repeated
cross-sectional study) conducted from January 2018 to January 2020 in
two Ministry of Health-affiliated hospitals in Mexico City: a
tertiary-level unit and a secondary-level unit. This study was approved
by the pertinent Institutional Review Board (IRB; Register
2010/010/3117). Written informed consent was obtained from each patient
prior to inclusion.
Women at different moments of spontaneous onset of the first stage of
labor were included. Women with multiple pregnancy, chronic diseases,
obstetric complications, and history of sexual intercourse or vaginal
medication within the last 48 hours were excluded. Women with clinical
or microbiological evidence of urinary tract infection or infectious
vulvovaginitis, as well as those with clinical or laboratory evidence of
premature rupture of membranes (PROM) and/or incomplete follow-up were
eliminated.
Obstetrics and Gynecology specialists, previously standardized,
evaluated the presence of clinical signs of spontaneous labor. Uterine
activity was recorded with a tocodynamometer. Cervical changes and
membrane integrity were assessed by vaginal examination. Gestational age
was determined by the date of the last menstrual period. Clinical and
demographic characteristics were obtained of all participants.
To represent initial changes associated to first phase of spontaneous
labor, pregnant women between 12 and 41 weeks of gestation (WoG) were
classified into one of the following five groups:
- Stage 0 : Women with no clinical evidence of myometrial
activity or cervical changes and intact fetal membranes. This stage
represents the total absence of labor.
- Stage 1 : Women with minimal sporadic uterine activity without
cervical changes and intact fetal membranes. None of these women gave
birth within posterior 10 weeks.
- Stage 2 : Women with self-perceived uterine contractions with
≤1 contraction event in 30 minutes, cervical softening and dilation
<2 cm, and intact fetal membranes. This stage is considered
as the initial or latent phase of spontaneous labor. Only women who
gave birth within posterior 24 hours after classification were
included in this group.
- Stage 3 : Women with painful uterine contractions ≤2 events in
30 minutes, cervical dilation 1-3 cm and intact fetal membranes.
Prelude phase of active labor. Only women delivering within posterior
12 hours after classification were included in this group.
- Stage 4 : Women with effective and regular uterine
contractions (three painful events in 10 minutes), cervix dilation
>3 cm and intact fetal membranes. This stage included
women in active labor. Only women delivering within posterior 6 hours
after classification were included in this group.
CVF samples were obtained by speculum examination. A Dacron tipped swab
was placed into the posterior fornix and transferred to transporting
buffer at 4ºC (0.05M Tris-base, 0.15M NaCl, 1% BSA, 0.1% Tween-20 and
protease inhibitors). Samples were centrifuged at 2000 x g at 4ºC for 15
minutes and supernatant fluid was collected and stored at -80ºC.
Cytokines in the CVF were measured with the Milliplex Human
Cytokine/Chemokine Magnetic Bead Panel and the MAGPIX Reader (Millipore,
Burlington, MA, USA); including: (1) chemokines: CXCL8 (IL-8 or
interleukin-8); (2) pro-inflammatory: interleukin-2 (IL-2),
interleukin-12p70 (IL-12p70), interferon-gamma (IFN-γ), tumor necrosis
factor alpha (TNF-α), interleukin-1 beta (IL-1β) and interleukin-6
(IL-6); (3) anti-inflammatory: interleukin-10 (IL-10), interleukin-4
(IL-4) and interleukin-1 receptor antagonist (IL-1RA).