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Plasma deoxyribonucleic acid concentrations of women in labor and umbilical cords
Plasma deoxyribonucleic acid concentrations of women in labor and umbilical cords RICHARD
C.
ALBERT
G.
Shreveport,
KAMM,
M.D.,
SMITH,
PH.D.
M.D.
Louisiana
The concentration of deoxyribonucleic acid (DNA) in plasma from 30 women in labor and plasma from the umbilical cords was determined with a rapid, inexpensive, and simple fluorometric method. These concentrations of DNA were compared to the plasma DNA concentrations of nonpregnant women. The mean plasma DNA concentration of women in labor was significantly lower than the mean plasma DNA concentration of nonpregnant women with the use of 99 per cent confidence limits. The mean plasma DNA concentration of umbilical cords during plasma DNA concentrations and of nonpregnant women
the third stage of labor was significantly lower of women in labor with the use of 99 per cent using 99.9 per cent confidence limits.
A RAPID, INEXPENSIVE, and simple fluorometric method for the determination of extracellular deoxyribonucleic acid (DNA) in human plasma has been developed,l making more extensive study in plasma DNA feasible. A study of the concentration of DNA in the plasma of women in labor and the differences in plasma DNA concentrations in umbilical cord blood compared to blood from the mother may provide information giving a more complete understanding of pregnancy and birth. Prior to the present study, very little work on plasma DNA in mothers and newborn infants has been done. Material
and
Received
methods
Revised Accepted
for
publication
May
8, 1974.
May
10,
February
mean limits
patient had no written clinic record or verbal history of illness prior to or during this pregnancy, approximately 7 ml. of whole blood was collected from a major vein of the forearm of the mother if she arrived at the labor unit in labor. Only patients in the first stage of labor were included in the study. If the expectant mother was not in labor, she was removed from the study. The whole blood was mixed with sodium ethylenediaminetetra-acetic acid (EDTA) as an anticoagulant and stored at room temperature (21 & 2O C. ) until a sample could be obtained from the umbilical cord at delivery. Blood stored for over 12 hours was discarded, and the mother was removed from the study. Those patients who had dystocia during labor or who required cesarean section or midforceps delivery were also removed from the study. Approximately 7 ml. of whole blood was collected by free flow from the umbilical cord during the third stage of labor. The sample was mixed with EDTA and then matched with the sample from the mother. The plasma from each sample was separated with low-speed centrifugation, and the paired samples were then analyzed for DNA concentration following digestion with ribonuclease with the fluorometric method described by Kamm and Smith.’ Kamm’ had previously shown that under the conditions utilized in this study the DNA concentration
All patients admitted to the labor unit of Confederate Memorial Medical Center, a universityaffiliated hospital, during the month of June, 1972, were initially entered into this study. Provided the From the Department of Pathololgy, Louisiana State University Medical Center, Confederate Memorial Medical Center.
than the confidence
and
5, 1974.
1974.
Reprint requests: Dr. Richard C. Kamm, Department of Pathology, Louisiana State University Medical Center, P. 0. Box 3932, Shreveport, Louisiana 71130. 29
30
Kamm
and
Smith
Table I. Comparison plasma
No. Mean S.D.
from
mothers
1
Plasma
86 13.8 2.8
of DNA concentrations in and from umbilical cords DNA
concentrations
30 12.2t 1.2
(mg./L.)
30
8.W D 1.9
with
the
use of
99.9
per
cent
confidence;
Comment
30 1.53 0.23
*Ratio of plasma DNA concentration of women in labor to plasma DNA concentration of their umbilical cords. tsignificantly different from the mean plasma DNA concentration of nonpregnant women at the 99 per cent confidence limit with the Mann-Whitney U test. $Significantly different from the mean plasma DNA concentration of women in labor at the 99 per cent confidence limit with the Mann-Whitney U test. SSignificantly different from the mean plasma DNA concentration of nonpregnant women at the 99.9 per cent
confidence limit with the Mann-Whitney
women limits.
U test.
remains unchanged for at least 5 days. It is felt that the difference in time between collection of the specimen from the mother and the collection from the umbilical cord did not affect the results obtained. Data obtained by Kamm and Smith’ from 86 female medical students and medical technologists who had no major illnesses within the past 5 years were used as a source for plasma DNA concentrations of nonpregnant women. The infants of the mothers included in the study were observed for 48 hours, and if an infant developed any difficulties or illnesses, the infant and the mother were excluded from the study. Results
A total of 30 mothers and their infants were included in this study. The plasma DNA concentration of the mother and infant were determined as described. The results are shown in Table I. It can be seen from Table I that the mean plasma DNA concentration of women in labor is significantly different from the mean plasma DNA concentration of nonpregnant women with the use of 99 per cent confidence limits and the Mann-Whitney U test.:’ Table I also demonstrates that the mean plasma DNA concentration of umbilical cords during the third stage of labor is significantly different from the mean plasma DNA concentration of women in labor with the use of 99 per cent confidence limits and from the mean plasma DNA concentration of nonpregnant
The plasma DNA concentrations of women in labor were found to be significantly lower than thr plasma DNA concentrations in nonpregnant women. Nordion and associates,q using the method of Schmidt and Tannhauser,: have previously studied the ribonucleic acid (RNA) concentrations in the sern of 32 pregnant women. They found the RNA concentration to be lower in women in the last trimester of pregnancy than during the second trimester. These two studies tend to imply that just prior to parturition there is a decline in nucleic acid concentrations in the concentrations of these nucleic acids in the plasma of nonpregnant women. If the nucleic acids are originating from cellular destruction, then parturition with its associated destruction of literally hundreds of cells would be expected to produce a rise in nucleic acid concentrations. However, this is not the case; therefore, simple cellular destruction cannot be presumed to be the source of the changes observed. There is a significantly lower concentration of extracellular DNA in the plasma of the umbilical cord as compared to the concentration in plasma from the mother. Nordion and associates,* studying serum, and Zaprudskaya and colleagues,” studying whole blood, have found that the KNA concentrations are likewise louver in the umbilical cord blood than in the mother’s blood. If the plasma concentration of DNA from the umbilical cord can be taken as an accurate indicator of the
Volume Number
121 1
Plasma
RNA phosphorus in the mother’s whole blood always exceed the amount in healthy children. They found the highest concentration of RNA in healthy children. They also observed that the RNA concentration rose in babies as they grew. Nordion and as-
DNA
of
women
in labor
and
umbilical
cords
31
sociates’ found that the RNA concentration in the serum from umbilical cords was about one half the RNA concentration in the serum drawn from the mother at the same time.
REFERENCES
1. 2. 3.
Kamm, 1972. Kamm, nessee Mann, 18:
R. C.,
and
Smith,
A. G.:
Clin.
Chem.
18: 519,
4.
R. C. : Doctoral dissertation, University of TenMedical Units, Memphis, Tennessee, 1970. H. B., and Whitney, D. R.: Ann. Math. Stat. 50, 1947.
5. 6.
Nordion, Biochim. Schmidt
S., Zinnari, A., Macagno, F., and Ferro M.: Appl. 13: 207, 1966. G., and Tannhauser, S. J.: J. Biol. Chem. 161: 83, 1945. Zaprudskaya, D. S., Loverdo, T. V., and Katuntseva, N. M.: Pediatriia 48: 28, 1969.
C.
ALBERT
G.
Shreveport,
KAMM,
M.D.,
SMITH,
PH.D.
M.D.
Louisiana
The concentration of deoxyribonucleic acid (DNA) in plasma from 30 women in labor and plasma from the umbilical cords was determined with a rapid, inexpensive, and simple fluorometric method. These concentrations of DNA were compared to the plasma DNA concentrations of nonpregnant women. The mean plasma DNA concentration of women in labor was significantly lower than the mean plasma DNA concentration of nonpregnant women with the use of 99 per cent confidence limits. The mean plasma DNA concentration of umbilical cords during plasma DNA concentrations and of nonpregnant women
the third stage of labor was significantly lower of women in labor with the use of 99 per cent using 99.9 per cent confidence limits.
A RAPID, INEXPENSIVE, and simple fluorometric method for the determination of extracellular deoxyribonucleic acid (DNA) in human plasma has been developed,l making more extensive study in plasma DNA feasible. A study of the concentration of DNA in the plasma of women in labor and the differences in plasma DNA concentrations in umbilical cord blood compared to blood from the mother may provide information giving a more complete understanding of pregnancy and birth. Prior to the present study, very little work on plasma DNA in mothers and newborn infants has been done. Material
and
Received
methods
Revised Accepted
for
publication
May
8, 1974.
May
10,
February
mean limits
patient had no written clinic record or verbal history of illness prior to or during this pregnancy, approximately 7 ml. of whole blood was collected from a major vein of the forearm of the mother if she arrived at the labor unit in labor. Only patients in the first stage of labor were included in the study. If the expectant mother was not in labor, she was removed from the study. The whole blood was mixed with sodium ethylenediaminetetra-acetic acid (EDTA) as an anticoagulant and stored at room temperature (21 & 2O C. ) until a sample could be obtained from the umbilical cord at delivery. Blood stored for over 12 hours was discarded, and the mother was removed from the study. Those patients who had dystocia during labor or who required cesarean section or midforceps delivery were also removed from the study. Approximately 7 ml. of whole blood was collected by free flow from the umbilical cord during the third stage of labor. The sample was mixed with EDTA and then matched with the sample from the mother. The plasma from each sample was separated with low-speed centrifugation, and the paired samples were then analyzed for DNA concentration following digestion with ribonuclease with the fluorometric method described by Kamm and Smith.’ Kamm’ had previously shown that under the conditions utilized in this study the DNA concentration
All patients admitted to the labor unit of Confederate Memorial Medical Center, a universityaffiliated hospital, during the month of June, 1972, were initially entered into this study. Provided the From the Department of Pathololgy, Louisiana State University Medical Center, Confederate Memorial Medical Center.
than the confidence
and
5, 1974.
1974.
Reprint requests: Dr. Richard C. Kamm, Department of Pathology, Louisiana State University Medical Center, P. 0. Box 3932, Shreveport, Louisiana 71130. 29
30
Kamm
and
Smith
Table I. Comparison plasma
No. Mean S.D.
from
mothers
1
Plasma
86 13.8 2.8
of DNA concentrations in and from umbilical cords DNA
concentrations
30 12.2t 1.2
(mg./L.)
30
8.W D 1.9
with
the
use of
99.9
per
cent
confidence;
Comment
30 1.53 0.23
*Ratio of plasma DNA concentration of women in labor to plasma DNA concentration of their umbilical cords. tsignificantly different from the mean plasma DNA concentration of nonpregnant women at the 99 per cent confidence limit with the Mann-Whitney U test. $Significantly different from the mean plasma DNA concentration of women in labor at the 99 per cent confidence limit with the Mann-Whitney U test. SSignificantly different from the mean plasma DNA concentration of nonpregnant women at the 99.9 per cent
confidence limit with the Mann-Whitney
women limits.
U test.
remains unchanged for at least 5 days. It is felt that the difference in time between collection of the specimen from the mother and the collection from the umbilical cord did not affect the results obtained. Data obtained by Kamm and Smith’ from 86 female medical students and medical technologists who had no major illnesses within the past 5 years were used as a source for plasma DNA concentrations of nonpregnant women. The infants of the mothers included in the study were observed for 48 hours, and if an infant developed any difficulties or illnesses, the infant and the mother were excluded from the study. Results
A total of 30 mothers and their infants were included in this study. The plasma DNA concentration of the mother and infant were determined as described. The results are shown in Table I. It can be seen from Table I that the mean plasma DNA concentration of women in labor is significantly different from the mean plasma DNA concentration of nonpregnant women with the use of 99 per cent confidence limits and the Mann-Whitney U test.:’ Table I also demonstrates that the mean plasma DNA concentration of umbilical cords during the third stage of labor is significantly different from the mean plasma DNA concentration of women in labor with the use of 99 per cent confidence limits and from the mean plasma DNA concentration of nonpregnant
The plasma DNA concentrations of women in labor were found to be significantly lower than thr plasma DNA concentrations in nonpregnant women. Nordion and associates,q using the method of Schmidt and Tannhauser,: have previously studied the ribonucleic acid (RNA) concentrations in the sern of 32 pregnant women. They found the RNA concentration to be lower in women in the last trimester of pregnancy than during the second trimester. These two studies tend to imply that just prior to parturition there is a decline in nucleic acid concentrations in the concentrations of these nucleic acids in the plasma of nonpregnant women. If the nucleic acids are originating from cellular destruction, then parturition with its associated destruction of literally hundreds of cells would be expected to produce a rise in nucleic acid concentrations. However, this is not the case; therefore, simple cellular destruction cannot be presumed to be the source of the changes observed. There is a significantly lower concentration of extracellular DNA in the plasma of the umbilical cord as compared to the concentration in plasma from the mother. Nordion and associates,* studying serum, and Zaprudskaya and colleagues,” studying whole blood, have found that the KNA concentrations are likewise louver in the umbilical cord blood than in the mother’s blood. If the plasma concentration of DNA from the umbilical cord can be taken as an accurate indicator of the
Volume Number
121 1
Plasma
RNA phosphorus in the mother’s whole blood always exceed the amount in healthy children. They found the highest concentration of RNA in healthy children. They also observed that the RNA concentration rose in babies as they grew. Nordion and as-
DNA
of
women
in labor
and
umbilical
cords
31
sociates’ found that the RNA concentration in the serum from umbilical cords was about one half the RNA concentration in the serum drawn from the mother at the same time.
REFERENCES
1. 2. 3.
Kamm, 1972. Kamm, nessee Mann, 18:
R. C.,
and
Smith,
A. G.:
Clin.
Chem.
18: 519,
4.
R. C. : Doctoral dissertation, University of TenMedical Units, Memphis, Tennessee, 1970. H. B., and Whitney, D. R.: Ann. Math. Stat. 50, 1947.
5. 6.
Nordion, Biochim. Schmidt
S., Zinnari, A., Macagno, F., and Ferro M.: Appl. 13: 207, 1966. G., and Tannhauser, S. J.: J. Biol. Chem. 161: 83, 1945. Zaprudskaya, D. S., Loverdo, T. V., and Katuntseva, N. M.: Pediatriia 48: 28, 1969.