Newer Predictors of Preeclampsia

Just a presentation covering most of the newer predictors of preeclampsia.

1. NEWER PREDICTORS OF PREECLAMPSIA DR ANUSHA RAO P PG II YR (OBG)

2. Traditional tests Novel predictors Roll-over test Isometric handgrip or cold pressor test Angiotensin-II infusion Midtrimester mean arterial pressure Platelet angiotensin-II binding Renin 24-hour ambulatory blood pressure monitoring Uterine artery or fetal transcranial Doppler velocimetry Human chorionic gonadotropin ( hCG ) Alpha-fetoprotein (AFP) Estriol Pregnancy- associated protein A (PAPP A) Inhibin A Activin A Placental protein 13 Corticotropin - releasing hormone Microtransferrinuria N -acetyl- 􏰂 - glucosaminidase Platelet count and activation Endothelial adhesion molecules

3. TRADITIONAL TESTS NOVEL PREDICTORS Serum uric acid Microalbuminuria Urinary calcium or kallikrein Fibronectin Prostaglandin Thromboxane C-reactive protein Cytokines Endothelin Neurokinin B Homocysteine Lipids Antiphospholipid antibodies Plasminogen activator-inhibitor (PAI)

4. NOVEL PREDICTORS Leptin P- selectin Angiogenic factors to include placental growth factor (PIGF), vascular endothelial growth factor (VEGF), fms -like tyrosine kinase receptor-1 (sFLT-1), Endoglin Antithrombin -III(AT-3) Atrial natriuretic peptide (ANP) 􏰂 2-microglobulin Genetic markers Free fetal DNA Serum proteonomic markers

5. The ideal biochemical marker for PE should exhibit the following characteristics: 1) Play a central role in the pathogenesis and be specific for the condition. 2) Appear early or before the clinical manifestations. Placental factors that can be detected early in pregnancy are likely to be good biochemical markers for PE prediction. However, placental disorders can cause IUGR without PE and vice versa, which makes the clinical evaluation of new markers particularly hard.

6. 3 ) Be easy and cheap to measure in maternal blood or urine. Few of the described factors are easy to measure; most of them require advanced laboratory system. 4) Show a high sensitivity and specificity . A small number of the described biochemical markers fulfill this requirement and strategies to use them in combination with other markers and/or, with PI measurements and other clinical parameters are being investigated. 5 ) Correlate with the severity of the condition . As the disease progresses, several organ systems are affected, which causes the number of factors to increase throughout pregnancy. A good candidate marker ought to appear early in pregnancy andcontinue to rise as the disease progresses. 6 ) Be non-detected or expressed at very low levels in normal pregnancies . Again, a placental factor is favored since the clinical symptoms disappear after removal of the placenta.

7. Angiotensin II sensitivity P reeclamptic women show an elevated pressor response to vasopressin. Although the performance of the test had a 77%sensitivity and 95% specificity in 1973, The largest most recent study of 494 healthy nulliparous women found a disappointing sensitivity of 22% with a specificity of 85% for prediction of pregnancy induced hypertensive disorders . The disappointing results of recent studies raises doubts about usefulness as a continuing test.

8. Renal laboratory tests

9. Urine albumin excretion rate  was the best predictive value for preeclampsia in hypertensive patient when blood for fibronectin , anthithrombin 3,alpha 1 microglobulin , U-N acetyl-beta- glucosominidase ,uric acid and albumin excretion rate was studied in 68 non pregnancy induced hypertensive and 40 chronic hypertensive, positive predictive value and specificity being 87.5 and 98.9 respectively

10. Elevated serum uric acid levels were associated with clinical severity of preeclampsia and perinatal outcomes. Despite a good amount of studies, not of all studies suggest that the serum uric acid levels may begin to rise before the onset of hypertension and proteinuria . Hyperuricaemia , an early sign of renal involvement in preeclampsia, is the result of reduced renal clearance due to altered tubular processing of uric acid preceding glomerular affliction, which will cause albminuria . However results from recent studies show that despite its long history of use, the discriminatory value of serum uric acid as a predictor of preeclampsia remains to be proven

11. Rodriques and Susuki showed that in women with a low levels of calcium/ creatinine ratio and high microalbuminuria , 84% developed PE. Microproteinuria levels above 375mg/l may be significant and thus could be utilized as a screening test for the early detection of a woman at risk of developing preeclampsia. However there is no diagnostic value of microalbuminuria and the calcium/ creatinine ratio when used alone.

12. When clinical utility of urinary soluble endoglin was compared with soluble fms -like tyrosine kinase 1 to placental growth factor (PIGF)ratio, soluble endoglin levels were significantly raised in preterm preeclampsia . However urinary endoglin has limitations to determine the severity of preeclampsia, and to differentiate between preeclampsia and chronic hypertension

13. Women with preeclampsia appear to present a unique urine proteomic fingerprint composed of SERPINA1 and albumin fragments that predicts preeclampsia in need of mandated delivery with highest accuracy . This characteristic proteomic profile also has the ability to distinguish preeclampsia from other hypertensive or proteinuric disorders in pregnancy.

14. Urine levels of inhibin A showed the greatest discrimination between severe preeclampsia and pregnant control women, when there was cut off of 45pg/mg. for urine creatinine . Women with greater than 90pg/mg urinary creatinine had a 17 fold relative risk of a clinically indicated delivery due to preeclampsia.

15. Measurement of urinary kallikrein (IUK)to creatinine ratio ( IUK:cr ) between 16-20 weeks of gestation for determining the risk of developing subsequent preeclampsia, with a sensitivity and specificity comparable to those reports as ascertained by other investigators using the well recognized, but less practical, angiotensin II sensitivity test

16. Urinary microtransferrinuria levels in pregnant women who subsequently developed preeclampsia and eclampsia where significantly higher than those pregnant women who remained normotensive. Microtranferrinuria as a predictor for preeclampsia had a sensitivity 93.5%,specificity 65%,positive predictive value of 83% and a negative predictive value of 98.4% Microtranferrinuria is potentially a more sensitive predictor of preeclampsia than microalbuminuria .

17. The urinary excretion of N-acetyl-beta- glucosaminidine , a lysosomal enzyme of the renal tubular cells, was increased in normal pregnant women and in woman with transient hypertension when compared to non pregnant healthy controls . In preeclampsia women, the increase was much higher than corresponding to their gestational age.

18. Maternal serum markers

19. Serum Inhibin –A was determined as a more sensitive marker for the prediction of preeclampsia than hCG . Addition of hCG data to inhibin did not improve the screening efficiency for preeclampsia suggesting that inhibin -A and hCG are markers of the same underlying pathological process.

20. Midtrimester beta- hCG levels alone correlated significantly with the severity of preeclampsia. However the combination of MShCG levels, body mass index(BMI),parity and age as a predictive test for preeclampsia was far superior to MShCG alone. This multi-factorial model could identify preeclampsia with a sensitivity of 70% and a specificity of 71 %.

21. Although the plasma TNF-alpha levels were higher in preeclamptics compared with normotensives,the levels cannot be used in the first, second trimester as a specific marker However they may be useful in the early third trimester.

22. Serum thrombomodulin antigen levels were measured in patients with preeclampsia, gestational hypertension and chronic hypertension . Levels were higher in preeclampsia than those with gestational hypertension or chronic hypertension. Thrombomodulin may serve as a clinical marker to differentiate preeclampsia from other disorders of pregnancy.

23. low levels of SHBG in the first half of pregnancy were reported as a promising early risk marker of the later development of preeclampsia. However as compared to normotensives controls, preeclamptic women exhibited no statically significant differences in the median levels of total testerone , free androgen index, sex hormone binding globulin or dehydroepiandrosterone .

24. Serum levels of collagen synthesis, procollagen I, carboxy -terminal peptide (PICP) and procollagen111 amino-terminal peptide (PIIIP), in patients with preeclampsia and controls. The markers were mildly elevated in preeclampsia, but unlikely to be useful in the prediction of preeclampsia.

25. Mid-trimester amniotic fluid cytokinase may reflect the maternal immune system in the maternal fetal interface and thus be predictive of preeclampsia. concentrations of interleukin (IL)- 6,8,10,11,12,15, tumor necrosis factor ( TNF )-alpha and transforming growth factor (TGF)-beta in the amniotic fluid at 14-16 weeks gestation from women with normal pregnancies and from those who subsequently developed severe preeclampsia are being studied.

26. Proteomics alterations in pre- eclampsia suggest that possible cellular battle against mitochondria-originated oxidative stress test resulting in recovery or apoptosis. The over expression of chaperonin 60, GST, VDAC, Erp29 and cathespin D in pre- eclampsia makes it a ideal marker of predicting preeclampsia

27. Serum CA125 variability. Lipoprotein a- Lp ( a) Coagulation Factors and Platelets Serum Oestriol Maternal serum alpha fetoprotein (MSAFP)

28. Pregnancy – associated protein A (PAPP-A) is a glycoprotein synthesized in the placenta and t he maternal plasma concentration increases through out pregnancy. PAPP-A has been used in combination with b-human chorionic gonadotropin (b- hCG ) and nuchal translucency thickness, to screen for trisomy 21, 13 and 18 at 11 to 13 weeks of gestation. In fetuses with normal chromosomes, decreased levels of PAPP-A in the 1 st trimester have been associated with increased risk for PE, IUGR, fetuses small for gestational age (SGA) and preterm delivery.

29. PAPP-A has been evaluated as a predictive and diagnostic biochemical marker for PE, but the screening performance, when used as a single biochemical marker, is only about 10 to 20 %. Combined with Doppler ultrasound, PAPP-A is a powerful predictive biochemical marker of PE with prediction rates of 70% at false positive rates of 5%. At term, plasma PAPP-A concentrations have been shown to increase in pregnancies complicated by PE and HELLP, but its concentration is still not predictive .

30. Recent reports suggest that free, extracellular fetal hemoglobin( HbF ) is involved in the pathogenesis of PE. Furthermore the heme and radical scavenger a1-microglobulin (A1M) is involved in the physiological defence against HbF . Their concentrations in maternal serum or plasma can be used as early predictive biochemical markers. Increased mRNA levels of HbF in the placental tissue and free HbF protein in the placental vascular lumen were described in women with PE .

31. Placental protein 13 (PP13 ) is a member of the galectin family and is produced by the placental trophoblast cells and is associated with normal placentation. In normal pregnancies, serum levels of PP13 slowly rise with gestational age. Several studies have shown lowered serum levels in the first trimester in pregnancies that subsequently developed PE.

32. When combining serum screening with Doppler ultrasound pulsatility index (PI), the prediction rate increased to 71% at a false positive rate of 10% . PP13 was concluded to be a reasonable biochemical marker for early onset and preterm PE but a weak marker for PE at term

33. Elevated levels of Soluble fms -like tyrosine kinase 1 (sFlt-1) occur before the clinical symptoms. The levels correlate with the time of onset of clinically manifest PE and partly with disease severity. Early-onset PE exhibits higher levels of sFlt-1. sFlt-1 increase is observed approximately 5 weeks before onset of PE.

34. Soluble Endoglin ( sEng ) truncated form of endoglin (CD105), a cell receptor for transforming growth factor-beta (TGF- β ), has been localized to both placental syncytiotrophoblasts and endothelial cells . Soluble endoglin is a second trimester marker for preeclampsia Circulating sFlt-1 and sEng may synergize and contribute to PE ,via different but additive mechanism, probably by inhibition of NO production, causing endothelial dysfunction. Circulating soluble endoglin levels were shown to increase 2-3 months before the onset of severe early-onset preeclampsia.

35. As a first trimester screening marker, soluble endoglin ( s- Eng ) shows conflicting results . Used in combination with Doppler ultrasound (PI) and PlGF , the prediction rate for early onset PE was 77.8% at a false positive rate of 5% .

36. sFlt-1 levels are stable during early & mid gestation ,then increase significantly during late stages. Angiogenic Markers In Healthy Pregnancy

37. Changes in circulating angiogenic factors from first to second trimester as predictors of PE Low or no increase in serum concentration of free PlGF , VEGF & high concentration of sFlt-1 a strong predictor of early PE. Low increase in PlGF & low increase in sFlt are associated with 10 fold higher risk of pre term PE. Low increase in PlGF in early pregnancy ,independent of change in sFlt-1 is associated with high risk –PE.

38. Mean sFlt-1 concentration and gestational age

39. Mean PIGF concentration and gestational age

40. Latest results :sFLT-1 free VEGF not useful in the 1st trimester screening Measurement of free-VEGF and sFlt-1 in maternal blood at 11 to 13 weeks of gestation is not useful in the prediction of pregnancies destined to develop PE.

41. sFlt-1 to PlGF Ratio as a Predictor for PE It is an index of antiangiogenic activity, that reflects changes in the balance between sFlt-1 & PlGF obviously seen in PE. Women with PE have reduced uteroplacental blood flow . Several studies have shown the predictive power of PlGF /sFlt-1 ratio from the second trimester . The prediction rate is about 89% . The transcription factor hypoxia –inducible factor 1 (HIF 1 alpha) regulates VEGF & Flt-1gene transcription. HIF-1 ,2 are elevated in pre eclamptic placenta.

42. Angiogenic Factors Prediction of PE In High Risk Women High risk women –Previous H/O PE, Multiple gestation, pre gestational DM, Chronic hypertension, chronic kidney disease, obesity & adolescent age. Study revealed that Rapid rise in the sFlt-1 to PlGF ratio with advancing gestation may be predictive of PE. Mean serum sFlt-1, sFlt-1/ PlGF were higher with early onset PE (< 34 weeks) Above ratio at 22-26 weeks was highly predictive of early onset PE. A 2 tiered screening approach may be a useful tool for prediction.

43. Twin pregnancies are associated with 2-3 fold increased risk for PE. Circulating levels of sFlt-1 PlGF ratio were twice as high as in singleton . But not accompanied by any change in levels of sFlt-1 mRNA & HIF – alpha protein, in the twin placentas, but were correlated with placental weight. Trisomy 13 a risk factor as gene for sFlt-1 is located on that chromosome. Smokers have low sFlt-1levels & less Incidence of PE.

44. Leptin , the product of the ob gene, is a hormone that is produced mostly in adipose cells . It is also produced in the placenta and may affect a number of processes in this organ, including angiogenesis, growth and immunomodulation. It has been suggested that leptin may be involved in the pathogenesis of preeclampsia. Many studies have reported high maternal leptin concentrations in the second trimester of pregnancy in women with preeclampsia. However , serum leptin concentrations in the early part of pregnancy did not differ between the preeclamptic and non- preeclamptic women.

45. Insulin-like growth factor-1 (IGF-1) is a hormone that may be involved in both normal and abnormal fetal growth. This hormone stimulates the renal and placental 1,25-dihydroxyvitamin D synthesis. It has been found that during preeclampsia the maternal IGF concentrations are lower compared with those in normal pregnancy.

46. It has been shown that there is a 5-fold increase in circulating fetal DNA concentrations in the maternal plasma with established preeclampsia compared with control pregnant subjects. This increase could be secondary to an increased entry of fetal cells, such as trophoblasts and erythroblasts, into the maternal circulation or the fetal DNA could liberate directly from dying cells in the placenta and a confirmation to this comes from the demonstration of widespread apoptosis in cytotrophoblasts obtained from the placental beds of preeclamptic pregnancies.

47. Neutrophil gelatinase associated lipocalin (NGAL), also known as lipocalin-2, siderocalin , uterocalin and 24p3, is a 25 kDa secreted protein that belongs to the family of lipocalins . NGAL is considered as the best and the earliest markers of acute kidney damage, where its presence can be detected in the urine within 2 hrs following the renal insult. A recent study demonstrated that the serum levels of NGAL increased at the end of the second trimester in women who subsequently developed pre- eclampsia compared to the control group

48. A uto-antibodies directed against the angiotensin-II type-1 receptor (AT1-AA) were detected in the blood from women with PE. The authors reporting this discovery provocatively suggested that plasma exchange could be a way of prevention Undoubtedly, these questions will have to be addressed before the clearance of AT1-AA can be considered as a therapeutic option. As a marker, AT1-AA appears to be less efficient than sFlt-1 .

49. Angiotensin receptor activating autoantibodies (AT1-AAs) may underlie many features of preeclampsia. AT1-AAs from preeclamptic patients activate angiotensin receptors (AT1R) on the surface of many cell types and may be responsible for many features of this serious pregnancy disorder. We have shown that antibody-induced receptor activation results in the mobilization of intracellular calcium and the activation of many genes. We propose that AT1-AAs activate AT1 receptors by promoting receptor homodimerization . ROS, reactive oxygen species. SMC, smooth muscle cells; EC, endothelial cells.

50. Cystatin C is a protease inhibitor widely used by clinicians as a sensitive marker for renal function and for estimation of glomerular filtration rate. The maternal plasma level of cystatin C is increased in women with PE and studies have demonstrated that the level of cystatin C is a reliable diagnostic marker for PE in the 1 st trimester.

51. 'Genes for pre- eclampsia ' discovered The US researchers from the Washington University School of Medicine in St. Louis analyzed DNA from over 300 pregnant women. 40 normal & remaining 250 were women who were being monitored for other health complications. Forty of these also went on to develop pre- eclampsia . DNA analysis revealed a few genetic errors shared by five of the 60 otherwise healthy women and seven of the 40 "higher-risk pregnancy" women who developed pre- eclampsia . The genes on which the errors were identified ( MCP factor I and factor H ) play a role in regulating immune response and the researchers believe this could explain their possible link to pre- eclampsia . Scientists have suspected that problems with the immune system provoke many cases of pre- eclampsia because women with lupus and certain other autoimmune diseases - like 250 of the women in the study - have an increased risk of the disorder. At best genes like these might identify 10-15% of pre- eclampsia , so it's relative importance may not be sensational. But it may allow us to study new treatments to prevent or delay the onset of pre- eclampsia and to know which women need closer surveillance.

52. Use of Doppler Pre - eclampsia screening in the first trimester of pregnancy: The combined use of transvaginal ultrasonography with the pulse-color Doppler technique allows the study of the uterine and umbilical circulation during the first trimester. Color Doppler imaging helps to identify the changes in uterine artery blood flow at 6 -9 weeks of gestation.

53. Doppler studies of brachial artery reactivity in women who have had pre- eclampsia show abnormal endothelial dependent flow-mediated arterial dilation three years after pregnancy. A relation between high resistance uterine artery waveforms in the second half of pregnancy and pre- eclampsia has already been established and persistent notching of the uterine artery Doppler waveform has shown promise as a screening test at 20 and 24 weeks.

54. Increased impedance to flow in the uterine arteries in both high-risk and low-risk pregnancies is associated with increased risk of development of pre- eclampsia and intrauterine growth restriction. Women with normal impedance to flow in the uterine arteries constitute a group that have a low risk of developing obstetric complications related to uteroplacental insufficiency.

55. Increased impedance to flow in the uterine arteries in pregnancy attending for routine antenatal care identifies about 50% of those that subsequently develop pre- eclampsia . Abnormal Doppler is better in predicting severe rather than mild pre- eclampsia . The sensitivity for severe pre- eclampsia is about 75%.

56. conclusion Screening for Down syndrome in the first trimester is a good example where a combination of ultrasound scanning and biochemical markers are used. Eg . It would be important to assess whether the measurement of other indices combined with inhibin A can produce a test with greater sensitivity for preeclampsia as occurs with triple or quadruple blood tests for Down's syndrome. This would open up new possibilities for the testing of preventive methods and the trials of treatments for this pregnancy-specific syndrome. Potential first trimester biochemical markers are PAPP-A , HbF and A1M . Both HbF and A1M play a role in the pathophysiology of PE . The biochemical markers appear as early as 10 weeks of gestation . Furthermore, they can be measured with basic ELISA techniques and show a high prediction rate at a low false positive level. Maternal plasma concentrations of free HbF have also been shown to correlate well with severity, i.e. blood pressure, in term PE pregnancies .

57. Angiogenic and anti- angiogenic factors are also very promising biochemical markers. Although the combination sFlt-1/ PlGF might not be useful in the first trimester, they are definitely well evaluated in the second trimester. Alterations of sFlt-1 and PlGF about 6 weeks before the onset of clinical symptoms and correlate with the severity of the disease. PlGF could be a promising biochemical marker even in the first trimester particularly if combined with HbF and A1M.

58. PP13 has shown potential as a biochemical marker of early onset PE, especially if combined with Doppler ultrasound uterine artery PI. However, as a general screening marker for all types of PE, the data is conflicting and needs further investigation. Restoration of angiogenesis balance in maternal circulation such as VEGF A 21 or neutralizing antibodies against sFlt-1 or sEnd may be the future therapy.

59. Preeclampsia may never be totally predictable, But better prediction would help to focus on antenatal care more effectively THANK YOU