Objective
We investigated mitochondrial DNA (mtDNA) content in the maternal circulation of normal pregnancies of different gestational ages and in pregnancies complicated by intrauterine growth restriction (IUGR).
Study Design
We examined 70 maternal blood samples: 13 nonpregnant women; 45 normal pregnancies, divided into the 3 trimesters; and 12 pregnancies complicated by IUGR. MtDNA content was determined by real-time quantitative polymerase chain reaction, using a genomic control and a target gene.
Results
A highly significant progressive reduction in circulating mtDNA was observed in pregnant women of first, second, and third trimesters and compared to nonpregnant women (mean value: 237, 188, 144, and 283, respectively; P < .001). Moreover, mtDNA was significantly increased in women carrying IUGR fetuses compared to women with normal pregnancies (430 vs 144; P < .001).
Conclusion
MtDNA could provide new insight into the mechanisms that occur during physiological gestation. Furthermore, mtDNA content may help recognize the IUGR disease in pregnancy.
Mitochondria are ubiquitous membrane-enclosed organelles recognized as the sites of aerobic oxidation of metabolic fuels. They contribute to many important functions including pyruvate and fatty acid oxidation, nitrogen metabolism, heme biosynthesis, and apoptosis regulation. Most notably, the mitochondrion is the site of the electron transport and oxidative phosphorylation that provides the bulk of cellular energy in the form of adenosine triphosphate.
Mitochondrial biogenesis requires a coordination of expression of the mitochondrial and the nuclear genomes and a constant cross talk between the 2. The communications between the mitochondria and the nucleus operate at 2 levels. One mechanism involves a set of transcription factors, or coactivators, in response to changes in environmental temperatures, external stimuli as changes in caloric intake or exercise, or changes in the levels of certain hormones. A second mechanism involves cellular responses to changes in the functional state of the mitochondria itself, a process also called “retrograde regulation.”
Some recent studies suggest the involvement of mitochondrial machinery influenced by external factors such as calorie restriction or temperature. Moreover, the regulation of mitochondrial biogenesis changes in hypoxic conditions of different organs or tissues. The involvement of relevant transcription factors that stimulate mitochondrial DNA (mtDNA) biogenesis and mtDNA maintenance has been postulated in the oxidative stress condition.
Pregnancy represents a serious challenge to the maternal body systems. The anatomical, physiological, and biochemical adaptations to pregnancy are profound. Many of these remarkable changes begin soon after fertilization and continue throughout gestation, and most occur in response to physiological stimuli provided by the conceptus and the placenta.
Intrauterine growth restriction (IUGR) constitutes an important clinical problem associated with increased perinatal morbidity and mortality, higher incidence of neurodevelopmental impairment, and increased risk of adult disease, such as diabetes and cardiovascular disease.
Placental vascular uneven perfusion together with altered transport function referred to as uteroplacental insufficiency has been demonstrated in IUGR.
Recently, we described higher mtDNA copies in IUGR placenta samples with a negative correlation to umbilical pO 2 levels. We hypothesized that this could result as an adaptation of the metabolic placental mechanism to the calorie restriction condition of the IUGR fetus, or as a compensatory mechanism of mitochondria to hypoxia.
The aim of this study was, first, to investigate whether mtDNA content in maternal blood changes during pregnancy. Second, we aimed to understand if the increased mtDNA copies number, which was found in IUGR placental tissues, should involve also maternal blood.
Materials and Methods
Patients
Patients were recruited at the Unit of Obstetrics and Gynecology, Department of Clinical Sciences “L.Sacco” and at the Ca’ Granda Ospedale Maggiore Policlinico, Milan, Italy. All pregnant and nonpregnant women were sampled on the occasion of a blood sample obtained for clinical reasons and informed that a small amount of blood was kept for the purpose of the study.
The study population consisted of 13 nonpregnant women, 45 normal pregnancies of different gestational ages (18 samples of first trimester, 16 samples of second trimester, and 11 samples of third trimester), and 12 pregnancies complicated by IUGR, which were diagnosed and treated in our department.
Nonpregnant women were of reproductive age. The inclusion criteria for the nonpregnant group were healthy patients with normal menstrual cycles, no previous gynecological problems, and no medication or hormonal contraceptive use.
All enrolled pregnancies were singleton. Exclusion criteria were gestational diabetes, abnormal fetal karyotype, fetal malformations, and infections. Gestational age was calculated from the last menstrual period and confirmed by routine ultrasonography at 11-12 weeks of gestation.
Normal pregnant women had physiological pregnancies and normal intrauterine fetal growth, confirmed by routine ultrasound scans performed at 20 and 32 weeks of gestation, according to Italian guidelines. The inclusion criteria for this group were: absence of maternal or fetal pathologies, no history of obstetric complications during pregnancies, and no pharmacologic treatment that could influence pregnancy outcome and fetal growth. All normal pregnant women delivered at term, between 37-42 weeks of gestation, an appropriate-for-gestational-age newborn according to birth weight references.
The inclusion criteria for IUGR were abdominal circumference measurements <10th percentile of reference values for fetuses of similar ages, together with a shift of the abdominal growth of >40th percentiles. Growth restriction was confirmed at birth by a neonatal weight <10th percentile according to Italian standards for birth weight and gestational age.
The exclusion criteria for both pregnant and nonpregnant women were maternal age >40 years, body mass index <18 or >25, past or present smoking, alcohol abuse, and drug addiction. Characteristics of the population are shown in Table 1 .
| Study population | NP (n = 13) | PI (n = 18) | PII (n = 16) | PIII (n = 11) | IUGR (n = 12) |
|---|---|---|---|---|---|
| Characteristics of population | |||||
| Maternal age, y | 34 ± 5 | 31 ± 5 | 33 ± 4 | 28 ± 5 | 34 ± 5 |
| Maternal BMI, kg/m 2 | 21 ± 2 | 21 ± 2 | 21 ± 2 | 21 ± 2 | 20 ± 1 |
| Gestational age at study, wk | h | 10 ± 2 | 19 ± 3 | 33 ± 5 | 33 ± 4 |
| Sampling results | |||||
| MtDNA content of maternal blood | 283 ± 50 | 237 ± 63 a , b , c | 188 ± 44 d , e | 144 ± 54 f | 430 ± 158 g |
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