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Biomedical subjects

I T Manyonda

Publications and source records attributed to I T Manyonda.

At least 19 recordsLinked to original sources
PubMed2001

Third consecutive complete posterior uterine rupture. A case report.

BACKGROUND: Management of pregnancy in a woman who has had a ruptured uterus on more than one occasion presents a great clinical and ethical challenge to the obstetrician. CASE: This appears to be the first report on complete uterine ruptures in three consecutive pregnancies. CONCLUSION: Prolonged hospital admission, intensive antenatal surveillance, antenatal steroid administration and elective premature delivery may give the best chance for a good outcome in these pregnancies. However, despite all these efforts, there is still a high risk of an unfavorable outcome.

Adult↗
PubMed2000

Excessive placental secretion of neurokinin B during the third trimester causes pre-eclampsia.

Pre-eclampsia is a principal cause of maternal morbidity and mortality, affecting 5-10% of first pregnancies worldwide. Manifestations include increased blood pressure, proteinuria, coagulopathy and peripheral and cerebral oedema. Although the aetiology and pathogenesis remain to be elucidated, the placenta is undoubtedly involved, as termination of pregnancy eradicates the disease. Here we have cloned a complementary DNA from human placental messenger RNA encoding a precursor protein of 121 amino acids which gives rise to a mature peptide identical to the neuropeptide neurokinin B (NKB) of other mammalian species. In female rats, concentrations of NKB several-fold above that of an animal 20 days into pregnancy caused substantial pressor activity. In human pregnancy, the expression of NKB was confined to the outer syncytiotrophoblast of the placenta, significant concentrations of NKB could be detected in plasma as early as week 9, and plasma concentrations of NKB were grossly elevated in pregnancy-induced hypertension and pre-eclampsia. We conclude that elevated levels of NKB in early pregnancy may be an indicator of hypertension and pre-eclampsia, and that treatment with certain neurokinin receptor antagonists may be useful in alleviating the symptoms.

Animals↗
PubMed2000

Efficient, rapid and reliable establishment of human trophoblast cell lines using poly-L-ornithine.

OBJECTIVE: Human trophoblast cells in primary culture are difficult to use for the rigorous study of trophoblast function because of contamination with other cell types, paucity of numbers, poor viability and inter-experiment variation engendered by the need to prepare fresh cells for each experiment. DNA-transfection to produce immortalized cells, or cells with extended life-span, has been the obvious approach to solve this problem. Although there have been a few reports in the literature describing trophoblast cell lines generated in this way, it is clear that to date the methods are difficult and few lines have been generated. The basic problem is that transfection efficiencies of different methods are cell type-specific. The objectives of this study were therefore to compare the transfection efficiencies of three commonly used techniques, to use the best technique to generate trophoblast cell lines, and to conduct preliminary characterization studies. METHODS: We have compared calcium phosphate co-precipitation, DEAE-dextran and poly-L-ornithine (PLO) DNA transfection protocols. We then used the most efficient to transfect human extravillous trophoblast with pSV3neo. RESULTS: Our modification of the PLO method has a transfection efficiency greater than 30 times that of the next best method. Several cell lines were established which had an extended life span and displayed an invasive phenotype, including the expression of MHC Class I framework antigens, human placental lactogen and human chorionic gonadotrophin, and thus have characteristics of extravillous trophoblast. In addition these cells express the integrin subunits b1, a1, and a3, all of which are known to be expressed in human trophoblast, and respond to IL-1alpha by increased secretion of GM-CSF. CONCLUSIONS: PLO is a highly efficient, rapid, reliable, simple and low-cost technique for the procurement of human trophoblast cell lines which retain most, if not all, the phenotype of the parental cell. These lines are potentially powerful tools in the rigorous study of trophoblast function.

Antigens, Polyomavirus Transforming↗
PubMed1998

Red cell distribution width (RDW) changes in pregnancy.

OBJECTIVE: As part of an exercise in establishing normograms of hematological parameters in pregnancy, we studied the red cell distribution width (RDW) in healthy pregnant women. METHODS: A longitudinal study of RDW measurements in 121 pregnant women at 16 and 34 weeks gestation and during labor and at Days 3 and 7 postpartum. All the women had uncomplicated pregnancies, minimum hemoglobin (Hb) of 11.0 g/dl at recruitment and took iron supplements from 16 weeks of gestation and until 7 days after delivery. All subjects went into spontaneous labor, 110 achieving a normal vaginal delivery while the remaining 11 were delivered by cesarean section. Two-way analysis of variance was used to study the changes in RDW between any given gestations to test the variability between and within subjects. RESULTS: RDW increased significantly (P < 0.0001) between 34 weeks of gestation and the onset of labor. No significant changes occurred between 16 and 34 weeks gestation, or during the 7 days postpartum. CONCLUSION: This is the first longitudinal study analyzing the between and within women changes in RDW with progression of pregnancy. The unexpected rise in the RDW during the last 4-6 weeks leading up to the onset of labor suggests increased bone marrow activity. The stimulus is unknown, but as RDW changes are highly significant there may well be a useful indicator of impending parturition.

Adolescent↗
PubMed1998

The phagocytic activity of human first trimester extravillous trophoblast.

It has been suggested previously that phagocytic activity in the human placenta is confined to cells of the macrophage lineage. However, earlier studies were hampered by the paucity and poor viability of cells inherent in primary trophoblast cell cultures, contamination by other cell types which themselves have phagocytic activity, lack of reliable markers of trophoblasts, and by limitations of methods available to demonstrate unequivocally the internalization of particulate material. We have overcome these limitations by using: (i) DNA transfection to provide unlimited supplies of pure trophoblast cell lines; (ii) human placental lactogen as a marker unique to trophoblast; and (iii) confocal microscopy to demonstrate unequivocally the intracellular locality of phagocytosed material. We found that both untransfected primary culture extravillous trophoblast cells, as well as the cell lines, had the capacity to phagocytose sheep red blood cells, Staphylococcus aureus and baker's yeast cells, and that this activity was inhibited by cytochalasin B and by culture at 4 degrees C. Phagocytic activity in trophoblast cells was less avid than that seen in a professional phagocyte. In physiological and pathological situations where tissue remodelling occurs, such as the rapid turnover in the periodontal ligament or during inflammation, epithelial cells and other cells that are not considered professional phagocytes actively phagocytose components of the extracellular matrix. We postulate that phagocytosis by human trophoblasts may play an important role in the extensive tissue remodelling that occurs during trophoblastic invasion of the decidua.

Animals↗
PubMed1998

A role for noradrenaline in pre-eclampsia: towards a unifying hypothesis for the pathophysiology.

OBJECTIVE: To compare plasma catecholamine (noradrenaline and adrenaline) levels in pre-eclamptic to normotensive pregnancy, and to study the activity of synthetic enzymes for catecholamines in placental and trophoblastic cell cultures. We postulated that catecholamines might be an important signal secreted by the fetoplacental unit in pre-eclampsia. METHODS: We recruited 12 women with pre-eclampsia and 12 pregnant women with nonproteinuric hypertension undergoing delivery by caesarean section, 23 normotensive women undergoing elective caesarean section at term, and 26 normotensive primigravid women with ongoing pregnancies at gestations equivalent to those women with pre-eclampsia. We measured venous blood concentrations of catecholamines. Following delivery, we studied tyrosine hydroxylase (the rate limiting enzyme for catecholamine synthesis) activity in placental tissue of these women as well as from four eclamptic women not in the observer study. We used Northern blot analysis to quantify mRNA for tyrosine hydroxylase and dopamine-beta-hydroxylase (D-beta-H, a non-rate-limiting synthetic enzyme for catecholamine) in placental tissue, as well as in trophoblast cells in primary culture and trophoblast cell lines. RESULTS: Venous blood concentrations of noradrenaline were significantly higher in pre-eclamptic women compared with normotensive women. Tyrosine hydroxylase activity was greater in placental tissue from pre-eclamptic and eclamptic compared with normotensive pregnancies, as were mRNA levels for this enzyme. The mRNA levels for the non-rate-limiting D-beta-H in women with pre-eclampsia were similar to those in normotensive pregnancies. First trimester trophoblast cells in primary culture and trophoblast cell lines transcript mRNA for tyrosine hydroxylase and D-beta-H. CONCLUSIONS: Trophoblasts have the capacity to secrete catecholamines, and we found increased activity of the rate-limiting synthetic enzyme in placental tissue from pre-eclamptic pregnancies. We postulate that the higher levels of catecholamines we found in the plasma of women with pre-eclampsia might be of placental origin. We hypothesise that in pre-eclampsia ischaemic trophoblast tissue secretes catecholamines as a physiological signal to increase maternal blood flow to the fetoplacental unit, which itself is spared the vasoconstrictor effects of catecholamines (placental vessels are known to be unresponsive to catecholamines). However, since the basic pathology--defective trophoblast invasion--is not corrected, the increased blood flow fails to resolve the ischaemia, and the secretion of catecholamines is therefore sustained or even enhanced. Noradrenaline is known to cause lipolysis. This results in breakdown of triglycerides to free fatty acids, which are oxidized to lipid peroxides. The latter are cytotoxic and cause widespread endothelial cell damage and dysfunction, culminating in the clinical syndrome of pre-eclampsia.

Adult↗
PubMed1993

Limiting dilution analysis of the allo-MHC anti-paternal cytotoxic T cell response. I: Normal primigravid and multiparous pregnancies.

Anti-paternal cytotoxic T lymphocyte precursor frequencies (CTLpF) were determined by limiting dilution analysis (LDA) in the peripheral blood of eight primigravid and seven multiparous women during the three trimesters of pregnancy. In five of these women the responses to cord blood lymphocytes (CBL) and paternal lymphocytes were also determined at the time of delivery and at 6 weeks post delivery. As controls, CTLpF against unrelated third party donors were determined. A wide range of CTLpF against all three groups of targets was found in both the primigravid and multiparous women, reflecting the wide range of frequencies found in random populations. These frequencies remained fairly constant during and 6 weeks after the pregnancy. Splitwell analysis demonstrated that the responses generated in our culture system were specific to the stimulator. The LDA data conform to single-hit kinetics, indicating that only cytotoxic T cells were limiting in the assay. Proliferative responses of maternal lymphocytes to paternal, cord blood and third party MHC antigens also remained unchanged as determined by time-course mixed lymphocyte reactions (MLR). Our data suggest that there is no significant allo-stimulation or suppression of the maternal immune system during normal pregnancy. The mother remains immunocompetent and is capable of both cytotoxic and proliferative responses to paternally-derived fetal MHC antigens. Our findings confirm that in normal pregnancy the trophoblast, which is devoid of classical MHC antigens, forms an effective immune barrier which prevents interaction of the maternal and fetal immune systems.

Cells, Cultured↗
PubMed1993

Limiting dilution analysis of the allo-MHC anti-paternal cytotoxic T cell response. II: Recurrent spontaneous abortion and the effect of immunotherapy.

Using limiting dilution analysis (LDA) we determined anti-paternal cytotoxic T lymphocyte precursor (CTLp) frequencies in the peripheral blood of 10 women with unexplained recurrent spontaneous abortion (RSA) before and after immunization with paternal lymphocytes. The women and their partners were HLA tissue-typed and none of the women had anti-paternal cytotoxic antibodies (APCA) before immunization. All other known causes of RSA were excluded. All 10 women were found to have high frequencies of specific anti-paternal cytotoxic T cells before immunization (range 1/1030 to 1/9574). Splitwell analysis showed that these cytotoxic cells were specific to paternal MHC antigens. These frequencies rose significantly following immunization (range 1/683 to 1/4652). The cytotoxic T lymphocyte frequencies against an HLA-mismatched third party varied from woman to woman, but were not affected by the immunization. The LDA data conformed to single-hit kinetics, indicating that only cytotoxic T cells were limiting in the assay. Our data are in sharp contrast to the previously held view that women with RSA may be hyporesponsive to paternal MHC antigens. Immunizing such women with paternal leucocytes further sensitizes them. These findings cannot be reconciled with a favourable outcome in the treatment of RSA with immunotherapy. We would argue that this treatment is at best of unproven value, and may even be harmful. That these women may sometimes have successful pregnancies following immunotherapy testifies to the effectiveness of the classical MHC antigen-deficient trophoblast as an immunological barrier between mother and fetus.

Abortion, Habitual↗
PubMed1992

A critical evaluation of the magnetic cell sorter and its use in the positive and negative selection of CD45RO+ cells.

In this paper, we report on our year-long experience with the magnetic cell sorter (MACS), and present a critical evaluation of its pitfalls and benefits. Satisfactory separation of lymphocytes into subsets with preservation of function can be achieved, but there are several drawbacks: in comparison with Dynal beads, MACS results in a higher cell loss due to the increased number of separation steps and requires depletion of plastic-adherent cells as these will engulf microbeads and contaminate the enriched fraction, and is more expensive. The advantage of MACS over Dynal beads is that the microbeads are biodegradable and do not interfere with proliferation assays: both the depleted and enriched fractions of cells can therefore be used in culture immediately following separation. We used MACS for the positive and negative selection of CD45RO cells: the enriched fraction was of high purity (greater than 98%), but a depleted fraction of greater than 90% purity could not be obtained even after running the same sample over three separating columns. Dynabeads, on the other hand, achieved 98% pure CD45RO-depleted fractions after three separation runs.

Antigens, CD↗