妊娠与饮酒：血液中磷脂酰乙醇水平与内源性中毒参数的关系

全文:

BACKGROUND

The development of normal pregnancy is accompanied by a series of adaptive responses aimed at maintaining dynamic homeostasis at all levels to ensure healthy fetal growth [1]. Functional changes in free radical homeostasis and neuroendocrine regulation play a significant role in this process, which may lead to alterations in middle molecule (MM) levels, serving as markers of endogenous intoxication [2]. MMs are biologically active peptide substances, including polyhydric alcohols, aminoglycans, and non-regulatory oligopeptides, depending on the type of diseases and complications; their concentrations increase with enhanced protein catabolism [3–5]. MMs are involved in microcirculatory disturbances, carbohydrate and energy metabolic disorders, and they also inhibit mitochondrial respiration and DNA synthesis. By exacerbating processes, they act as secondary toxins and can be considered prognostic factors of metabolic disorders [6]. MMs have diverse origins: alimentary (from food), from intestinal microbiota metabolites, and endogenous (protein proteolysis products). Due to their varied biological activities—disruption of biomembrane ion permeability, inhibition of enzymatic systems, and binding of essential proteins—they contribute significantly to endotoxicosis [7, 8].

Endotoxicosis is a complex and multifaceted pathophysiological process that profoundly affects the mother–placenta–fetus system during gestation. Pregnancy induces hemodynamic adaptations and alters the lipid profile with the predominance of unsaturated fatty acids and cholesterol, potentially aggravating endogenous intoxication. One of the key pathophysiological mechanisms underlying endotoxicosis is the activation of lipid peroxidation processes triggered by oxygen radicals, along with increased phospholipase system activity. Progression of endotoxicosis in pregnant women is further promoted by hormonal imbalance, diabetes mellitus, liver diseases, and impaired detoxification systems [9, 10]. These disturbances may occur in the context of maternal alcohol consumption. Ethanol, even at low doses, exerts detrimental effects on the embryo. Several fetal neurophysiological pathologies induced by ethanol are now collectively recognized as fetal alcohol syndrome, observed in approximately half of children born to women with alcohol dependence. During pregnancy, ethanol exposure may lead to oligohydramnios, placental aging, severe toxicosis, and accumulation of toxic metabolites in the amniotic fluid [11]. It has been shown that levels of lipid peroxidation intermediates are significantly higher in alcohol-consuming women compared with controls [12].

It is believed that MMs can penetrate the placental barrier and exert adverse effects on the fetus, causing various multiorgan disturbances [13]. The risk of impaired development of the fetus and placenta increases during critical periods of pregnancy because of hormonal insufficiency (6–12 weeks), the development of isthmic–cervical insufficiency, as well as gestational diabetes mellitus and pre-eclampsia (18–22 weeks), late gestosis, placental insufficiency, and premature placental abruption (28–32 weeks). Therefore, it is of particular interest to assess the levels of these factors at these specific time points. In addition, from week 38, the maternal organism undergoes intensive preparation for the upcoming delivery; this period is extremely important because fetal development is fully completed [14].

Known alcohol biomarkers are considerably variable in sensitivity and specificity [15]. The most reliable marker for the detection of ethanol intoxication is phosphatidylethanol (PEth), the determination of which in biological fluids is considered a promising method for diagnosing episodic alcohol intake and chronic alcohol intoxication. PEth is a glycerophospholipid formed from phosphatidylcholine in various tissues in the presence of ethanol. Because of its long half-life after alcohol consumption, PEth accumulates in the blood and can be detected for up to 28 days after alcohol intake [16, 17]. To date, 48 PEth homologues have been identified, among which PEth 16:0/18:1 is the most informative [18].

The study aimed to assess endogenous intoxication by the concentration of MMs in venous blood of women depending on serum phosphatidylethanol levels at different stages of pregnancy.

MATERIALS AND METHODS

The study was conducted from 2021 to 2024 in accordance with the Declaration of Helsinki of the World Medical Association (1964, amended 2013) and was approved by the Biomedical Ethics Committee of the Scientific Center for Family Health and Human Reproduction (meeting extract No. 2 of March 4, 2021). The written informed consent was obtained from all participants.

A prospective study included 163 pregnant women receiving care at Irkutsk City Clinical Hospital No. 8. The sample size was not pre-determined. Inclusion criteria: current pregnancy; age 18–40 years; signed informed consent; regular follow-up at the medical facility. Non-inclusion criteria: HIV infection; viral hepatitis; diabetes mellitus; exacerbation of chronic diseases; acute respiratory viral infection; COVID-19; participation in an in vitro fertilization program; sexually transmitted infections; arterial hypertension. Exclusion criteria: withdrawal of consent, protocol violations, technical problems with samples, gestational diabetes mellitus, pre-eclampsia, and thyrotoxicosis. The parameters were evaluated according to gestational age at 6–12, 18–22, 28–32, and 38–40 weeks of pregnancy. The exclusion criteria and deliveries before 38 weeks of gestation resulted in a subset of women unable to provide biological samples at all four time points.

The group distribution was as follows:

• 6–12 weeks of gestation: group 1, n = 62; group 2, n = 66; group 3, n = 35;
• 18–22 weeks of gestation: group 1, n = 53; group 2, n = 57; group 3, n = 35;
• 28–32 weeks of gestation: group 1, n = 43; group 2, n = 49; group 3, n = 28;
• 38–40 weeks of gestation: group 1, n = 34; group 2, n = 35; group 3, n =

Based on PEth concentrations, groups of women were identified according to alcohol consumption levels: group 1 with PEth ≤8 ng/mL (non-drinkers, control); group 2 with 8–45 ng/mL (drinking less than one dose); group 3 with ≥45 ng/mL (drinking more than one dose) [12].

Fasting blood samples were collected from the median cubital vein according to standard procedures. The quantitative determination of the direct alcohol biomarker PEth 16:0/18:1 in plasma was performed using High-Performance Liquid Chromatography–Mass Spectrometry on Shimadzu LCMS-8060 system (Japan). The validated lower limit of quantification for PEth 16:0/18:1 was 1 ng/mL.

MM levels were measured at four wavelengths (238, 254, 260, and 280 nm) using SF-2000 spectrophotometer (Russia) [19]. MM fractions were expressed in arbitrary units (AU) of optical density. Distribution coefficients (238/260, 238/280, 280/254) were also calculated.

Statistical analysis was performed using STATISTICA 10.0 software (StatSoft Inc, USA). The normality of distribution for continuous variables was assessed with the Lilliefors-corrected Kolmogorov–Smirnov test and the Shapiro–Wilk test. Because the data were normally or near-normally distributed, results were presented as means and standard deviations (m ± SD). Intergroup differences were assessed using the parametric Student t-test with Bonferroni correction; differences were considered statistically significant at p ≤ 0.017.

RESULTS

MM levels at different stages of pregnancy in the study groups are shown in Table 1. At 28–32 weeks, a decrease in MM levels (λ = 238 nm) was observed in alcohol-consuming pregnant women compared with controls, regardless of PEth blood levels (p = 0.013 and p = 0.003 in groups 2 and 3, respectively). Before delivery, MM levels were lower in group 3 compared with controls (p = 0.004). A significant decrease in MM levels (λ = 280 nm) was detected in group 3 compared with group 2 (p = 0.017).

 

Table 1. Level of middle molecules (MM) at different stages of pregnancy in the studied groups of women (M±SD)

Gestational age	Group 1	Group 2	Group 3	Significance level
MM238
6–12 weeks	0.30±0.21	0.27±0.12	0.24±0.16	–
18–22 weeks	0.27±0.14	0.29±0.15	0.29±0.13	–
28–32 weeks	0.34±0.17	0.26±0.13	0.22±0.12	р1–2=0.013 р1–3=0.003
38–40 weeks	0.33±0.16	0.27±0.16	0.19±0.09	р1–3=0.004
MM254
6–12 weeks	0.23±0.16	0.19±0.09	0.21±0.12	–
18–22 weeks	0.21±0.09	0.22±0.09	0.22±0.10	–
28–32 weeks	0.23±0.11	0.20±0.07	0.20±0.07	–
38–40 weeks	0.23±0.10	0.21±0.07	0.18±0.04	–
MM260
6–12 weeks	0.22±0.10	0.21±0.08	0.22±0.12	–
18–22 weeks	0.25±0.29	0.23±0.09	0.23±0.10	–
28–32 weeks	0.23±0.11	0.22±0.06	0.21±0.07	–
38–40 weeks	0.23±0.11	0.23±0.06	0.19±0.05	–
MM280
6–12 weeks	0.28±0.12	0.25±0.08	0.27±0.13	–
18–22 weeks	0.26±0.10	0.29±0.10	0.29±0.10	–
28–32 weeks	0.30±0.13	0.29±0.10	0.28±0.08	–
38–40 weeks	0.31±0.12	0.30±0.07	0.24±0.06	р2–3=0.017

 

The calculation of distribution coefficients showed that, compared with controls, alcohol-consuming women, regardless of the PEth 16:0/18:1 concentration, had significantly lower values of the 238/260 distribution coefficient (p = 0.007 and p < 0.001 in Groups 2 and 3, respectively) and the 238/280 distribution coefficient (p = 0.002 and p < 0.001 in Groups 2 and 3, respectively) at 28–32 weeks of gestation (Table 2). Before delivery, decreases in these coefficients were noted only in group 2 (p = 0.006 for 238/260; p = 0.015 for 238/280). The 280/254 distribution coefficient was higher in alcohol-consuming women at 28–32 weeks compared with controls (p = 0.003 and p = 0.014 in Groups 2 and 3, respectively).

 

Table 2. Distribution coefficients at different stages of pregnancy in the studied groups of women (M±SD)

Gestational age	Group 1	Group 2	Group 3	Significance level
238/260
6–12 weeks	1.38±0.72	1.41±0.90	1.09±0.66	–
18–22 weeks	1.31±0.62	1.27±0.54	1.29±0.54	–
28–32 weeks	1.48±0.62	1.15±0.54	0.99±0.47	р1–2=0.007 р1–3 <0.001
38–40 weeks	1.51±0.63	1.13±0.48	1.04±0.57	р1–2=0.006
238/280
6–12 weeks	1.11±0.64	1.18±0.98	0.87±0.50	–
18–22 weeks	1.07±0.52	1.02±0.43	1.01±0.39	–
28–32 weeks	1.15±0.47	0.86±0.41	0.76±0.38	р1–2=0.002 р1–3 <0.001
38–40 weeks	1.12±0.47	0.86±0.39	0.82±0.44	р1–2=0.015
280/254
6–12 weeks	1.33±0.29	1.34±0.27	1.37±0.25	–
18–22 weeks	1.30±0.22	1.38±0.27	1.35±0.23	–
28–32 weeks	1.33±0.21	1.48±0.25	1.47±0.25	р1–2=0.003 р1–3=0.014
38–40 weeks	1.41±0.27	1.45±0.19	1.39±0.16	–

 

DISCUSSION

The mechanisms of biochemical adaptation in normal pregnancy and in obstetric diseases involve changes in the placental metabolic regulation. The development of pregnancy are characterized by improved oxygen delivery to the fetus and placenta due to thinning of the placental membrane, which facilitates the transfer of MMs across the placental barrier and their effects on the fetus [20, 21].

More intensive catabolic processes in the third trimester of normal pregnancy, compared with alcohol-consuming women, are indicated by higher levels of peptide substances absorbing at 238 nm. The increased production of reactive oxygen species occurs due to the enhanced metabolism, high oxygen consumption, and fatty acid utilization. During the third trimester, insulin resistance increases, fat catabolism and release of free fatty acids intensify, leading to the increased hydrogen peroxide production [22]. The MM238 includes catabolic products, natural degradation products of cells and tissues, and microbial particles [23]. Lower values of these parameters in the third trimester in alcohol-consuming pregnant women may indicate lower catabolic activity and immunogenesis during this period, which, together with reduced anabolic intensity, adversely affects the mother–placenta–fetus system.

It is known that MM280 reflects the anabolic pool and demonstrates activation of reparative and synthetic processes in cells and tissues. At this wavelength, phenols, tyrosine, tryptophan, and phenylalanine—important for neuronal processes—have maximal absorption. Several studies have demonstrated the impaired synthesis and metabolism of aromatic amino acids with alcohol consumption. Altered metabolism of these amino acids leads to central nervous system neurotransmitter imbalance, which underlies psychiatric and neurological disorders in alcoholism [24].

Endogenous intoxication may result both from elevated concentrations of certain substances and from the disruption of equilibrium between components of homeostatic processes. For this reason, calculation of coefficients provides an additional meaningful characteristic of the development and severity of pathologic processes. The peptide-to-nucleotide coefficient at 238/260 nm reflects the ratio of peptide content shifts, whereas the aromaticity coefficient at 238/280 nm reflects the ratio of aromatic to nonaromatic chromophores [25].

A decrease in aromaticity coefficient with alcohol consumption, regardless of dose, beginning at 28 weeks of gestation may indicate increased cellular synthesis of biologically active compounds promoting labor activity and potentially contributing to pre-term birth. A decrease in the peptide-to-nucleotide coefficient in these groups suggests accumulation of toxic metabolic products (incomplete protein breakdown products and hydrophobic toxins), which may result from consumption of ethanol-containing products.

The ratio of the 280/254 fractions in Groups 2 and 3 was higher than in controls. An increase in this parameter may indicate intensified catabolic processes, more active lipid peroxidation, and enhanced immunogenesis. Previous studies have shown that even minimal alcohol intake in the first trimester of pregnancy activates lipid peroxidation processes [12].

The processes occurring during ethanol metabolism promote oxidation of proteins and lipids, DNA destruction, and mitochondrial dysfunction, all of which ultimately lead to apoptosis and cellular injury. The disruption of key placental functions occurs as a result of destructive and proliferative changes induced by the toxic effects of alcohol. Chronic alcohol intoxication leads to reduced placental mass and dystrophic and necrotic changes in the chorionic epithelium, which may subsequently manifest as chronic fetoplacental insufficiency, fetal hypoxia, and growth restriction [26, 27].

Elevated levels of progesterone, cortisol, prolactin, estradiol, and other hormones are characteristic of the hormonal shift during gestation [28], which, in turn, may intensify free radical oxidation processes [29, 30] and alter the pool of middle-mass peptides. It has been shown that the harmful effects of ethanol involve stimulation of the hypothalamic–pituitary–adrenal system, with the potential for overstrain and failure of compensatory mechanisms, leading to imbalance in redox status [31]. In addition, women with the laboratory-confirmed alcohol consumption in early gestation are significantly more likely to experience intrauterine growth restriction, anemia, and pre-term birth [11], which may be associated with the involvement of imbalance in the prooxidant–antioxidant system and alterations in the pool of middle-molecular peptides.

This study has several limitations. First, not all pregnant women were able to provide biological material for analysis at all gestational time points because of the exclusion criteria. Second, alcohol consumption by each woman at all stages of pregnancy was not assessed, which precluded longitudinal observation of changes in middle-molecular fractions across gestational ages in the same participant. Future studies are planned to define groups of women and investigate changes in MM concentrations longitudinally, depending on alcohol consumption in different trimesters of pregnancy.

CONCLUSION

The findings indicate reduced levels of specific fractions of middle-molecular toxins reflecting both anabolic and catabolic pools in alcohol-consuming women, which may be associated with serious metabolic disturbances in the mother–placenta–fetus system. Distribution coefficients proved to be sensitive markers for monitoring endogenous intoxication in pregnant women, suggesting a predominance of catabolic processes with accumulation of catabolic products and a possible increased risk of pre-term delivery, regardless of alcohol dose.

The problem of alcohol use in the modern stage of social development remains one of the most urgent issues, the resolution of which may reduce the risk of gestational complications and fetal malformations. Monitoring the levels of middle-molecular toxin fractions may help in decision-making regarding pharmacologic correction or educational interventions for pregnant women.

ADDITIONAL INFORMATION

Author contributions: N.V. Semenova: conceptualization, methodology, formal analysis, writing—original draft; A.N. Karacheva, T.A. Bairova, A.V. Belskikh, E.V. Belyaeva, A.Yu. Sambyalova, O.A. Ershova, N.V. Protopopova: investigation; E.A. Novikova: source search and analysis; O.A. Nikitina: data curation, formal analysis; A.Yu. Marianian: methodology; S.I. Kolesnikov, L.I. Kolesnikova: writing—review & editing. All the authors confirm that their authorship meets the ICMJE criteria (all authors made substantial contributions to the conceptualization, investigation, and manuscript preparation, and reviewed and approved the final version prior to publication).

Ethics approval: The study was approved by the Local Ethics Committee of the Federal State Budgetary Scientific Institution Scientific Center for Family Health and Human Reproduction Problems (protocol No. 2 dated March 4, 2021).

Consent for publication: All participants provided written informed consent prior to inclusion in the study.

Funding sources: The study was carried out under research project No. 123051600012-7.

Disclosure of interests: The authors have no relationships, activities, or interests for the last three years related to for-profit or not-for-profit third parties whose interests may be affected by the content of the article.

Statement of originality: No previously published material (text, images, or data) was used in this work.

Data availability statement: The editorial policy regarding data sharing does not apply to this work, as no new data was collected or created.

Generative AI: No generative artificial intelligence technologies were used to prepare this article.

Provenance and peer-review: This paper was submitted unsolicited and reviewed following the standard procedure. The peer review process involved two external reviewers, a member of the editorial board, and the in-house scientific editor.

作者简介

Natalya V. Semenova

Scientific Сentre for Family Health and Human Reproduction Problems

编辑信件的主要联系方式.
Email: natkor_84@mail.ru
ORCID iD: 0000-0002-6512-1335
SPIN 代码: 6606-0160

Dr. Sci. (Biology)

俄罗斯联邦, Irkutsk

Elizaveta A. Novikova

Scientific Сentre for Family Health and Human Reproduction Problems

Email: yelizaveta_novikova_2001@bk.ru
ORCID iD: 0009-0001-1207-3309
SPIN 代码: 2734-2998
俄罗斯联邦, Irkutsk

Olga A. Nikitina

Scientific Сentre for Family Health and Human Reproduction Problems

Email: olga_tolpygina@mail.ru
ORCID iD: 0000-0003-1926-9694
SPIN 代码: 1799-4973

Cand. Sci. (Biology)

俄罗斯联邦, Irkutsk

Anait Yu. Marianian

Scientific Сentre for Family Health and Human Reproduction Problems

Email: anait_24@mail.ru
ORCID iD: 0000-0002-9544-2172
SPIN 代码: 2584-6861

MD, Dr. Sci. (Medicine)

俄罗斯联邦, Irkutsk

Anastasia N. Karacheva

Scientific Сentre for Family Health and Human Reproduction Problems

Email: kalkova_nastya@mail.ru
ORCID iD: 0000-0002-7626-020X
SPIN 代码: 3082-0796
俄罗斯联邦, Irkutsk

Sergey I. Kolesnikov

Scientific Сentre for Family Health and Human Reproduction Problems

Email: iphr@sbamsr.irk.ru
ORCID iD: 0000-0003-2124-6328
SPIN 代码: 1752-6695

MD, Dr. Sci. (Medicine), Professor, Academician of the Russian Academy of Sciences

俄罗斯联邦, Irkutsk

Tatyana A. Bairova

Scientific Сentre for Family Health and Human Reproduction Problems

Email: tbairova38@mail.ru
ORCID iD: 0000-0003-3704-830X
SPIN 代码: 2039-3728

MD, Dr. Sci. (Medicine)

俄罗斯联邦, Irkutsk

Aleksey V. Belskikh

Scientific Сentre for Family Health and Human Reproduction Problems

Email: alex590750@yandex.ru
ORCID iD: 0000-0003-3678-7274
SPIN 代码: 2004-1578

Cand. Sci. (Chemistry)

俄罗斯联邦, Irkutsk

Elena V. Belyaeva

Scientific Сentre for Family Health and Human Reproduction Problems

Email: belyeva_irk@mail.ru
ORCID iD: 0000-0001-6050-5287
SPIN 代码: 7865-3241

Cand. Sci. (Biology)

俄罗斯联邦, Irkutsk

Alexandra Yu. Sambyalova

Scientific Сentre for Family Health and Human Reproduction Problems

Email: sambialova95@mail.ru
ORCID iD: 0000-0001-5790-6282
SPIN 代码: 3435-5028
俄罗斯联邦, Irkutsk

Oksana A. Ershova

Scientific Сentre for Family Health and Human Reproduction Problems

Email: oksana111088@mail.ru
ORCID iD: 0000-0003-0690-4636
SPIN 代码: 3002-7757

Cand. Sci. (Biology)

俄罗斯联邦, Irkutsk

Natalia V. Protopopova

Scientific Сentre for Family Health and Human Reproduction Problems

Email: doc_protopopova@mail.ru
ORCID iD: 0000-0002-1740-228X
SPIN 代码: 7622-3480

MD, Dr. Sci. (Medicine), Professor

俄罗斯联邦, Irkutsk

Lubov I. Kolesnikova

Scientific Сentre for Family Health and Human Reproduction Problems

Email: iphr@sbamsr.irk.ru
ORCID iD: 0000-0003-3354-2992
SPIN 代码: 1584-0281

MD, Dr. Sci. (Medicine), Professor, Academician of the Russian Academy of Sciences

俄罗斯联邦, Irkutsk

参考

补充文件