Predicting Intrauterine Transfusion Interval and Perinatal Outcomes in Alloimmunized Pregnancies: Time-to-Event Survival Analysis.

BACKGROUND: The risk factors determining the frequency of intrauterine transfusions (IUTs) for severely affected red blood cell alloimmunized singleton pregnancies are not well known. OBJECTIVE: To assess factors associated with IUT frequency and adverse pregnancy outcomes in transfused pregnancies. METHODS: Retrospective cohort analysis of 246 consecutive cases between 1991 and 2014. Time-to-event survival analysis for repeated events was used to evaluate risk of subsequent IUT. Multivariable logistic regression assessed odds of a composite adverse pregnancy outcome (intrauterine fetal death, termination of pregnancy, neonatal death, preterm birth <34 weeks' gestation). RESULTS: Full information was available on232 cases (94.3%) and 716 IUTs. Fetal hydrops was associated with increased frequency (hazard ratio [HR] 1.29 [95% CIs 1.15-1.47, p < 0.001]) while higher fetal hemoglobin (Hb) pre-IUT (HR) 0.99 (95% CI 0.99-1.00, p = 0.021) and post-IUT (HR 0.99 [95% CI 0.99-1.00] p = 0.042), and higher transfused blood volume (HR 0.98 [95% CI 0.97-0.99] p < 0.001) were associated with reduced IUT frequency. Adverse pregnancy outcomes were more likely with lower gestational age (GA) at initial IUT. Antibody type was not associated with IUT frequency or adverse pregnancy outcomes. CONCLUSIONS: Hydrops is associated with increased IUT frequency while lower GA at initial IUT is associated with higher adverse pregnancy outcomes in alloimmunized pregnancies.Higher transfused blood volumes, pre- and post-IUT Hb are associated with lower IUT frequency.

The near disappearance of fetal hydrops in relation to current state-of-the-art management of red cell alloimmunization.

OBJECTIVE: In this study, we aim to evaluate trends in the condition of fetuses and neonates with hemolytic disease at the time of first intrauterine transfusion (IUT) and at birth, in relation to routine first-trimester antibody screening, referral guidelines, and centralization of fetal therapy. METHOD: We conducted a 30-year cohort study including all women and fetuses treated with IUT for red cell alloimmunization at the Dutch national referral center for fetal therapy. RESULTS: Six hundred forty-five fetuses received 1852 transfusions between 1 January 1987 and 31 December 2016. After the introduction of routine first-trimester antibody screening, the hydrops rate declined from 39% to 15% (OR 0.284, 95% CI, 0.19-0.42, P < 0.001). In the last time cohort, only one fetus presented with severe hydrops (OR 0.482, 95% CI, 0.38-0.62, P < 0.001). Infants are born less often <32 weeks (OR 0.572, 95% CI, 0.39-0.83, P = 0.004) and with higher neonatal hemoglobin (P < 0.001). Neonatal hemoglobin was positively independently associated with gestational age at birth, fetal hemoglobin, and additional intraperitoneal transfusion at last IUT. CONCLUSION: Severe alloimmune hydrops, a formerly often lethal condition, has practically disappeared, most likely as a result of the introduction of routine early alloantibody screening, use of national guidelines, and pooling of expertise in national reference laboratories and a referral center for fetal therapy.

Production and characterization of monoclonal antibodies against α-globin chain-containing human hemoglobins for detecting α-thalassemia disease.

Monoclonal antibodies against α-globin containing human Hbs, named AMS-Alpha1 and AMS-Alpha 2, were produced by the hybridoma technique using spleen cells enriched by the newly developed B lymphocyte enrichment protocol. These two monoclonal antibodies were of IgM class, reacting to only intact form of human Hbs A, A2, E, and F, which contain α-globin chain. By the indirect ELISA, the AMS-Alpha1 and AMS-Alpha 2 quantified less amount of α-globin chain containing hemoglobins in HbH disease than the SEA-α thalassemia 1 carriers and normal individuals. It was thus anticipated that these monoclonal antibodies can be used for detecting Hb Bart's hydrops fetalis in which no α-globin chain is produced.

Management of fetal hydrops due to maternal antibodies against a low incidence paternal red cell antigen: a novel insight from clinical practice.

A rare case of a low incidence red cell antigen causing severe fetal allo-immune red cell disease is presented. Discussion of how this can be diagnosed and successfully managed antenatally using middle cerebral artery Doppler ultrasound and maternal antibody titre levels for fetal surveillance and timing of intervention with intrauterine transfusion.

Exchange transfusion of least incompatible blood for severe hemolytic disease of the newborn due to anti-Rh17.

HDN attributed to the rare Rh variants has become more and more significant caused by anti-D, but the compatible blood is usually very difficult to obtain when exchange transfusion is required. We treated a 10-hour neonate of O, D + C + c - E - e+ blood group with severe HDN due to anti-Rh17 with least incompatible blood typed O, D + C - c + E + e-. The neonatal hemolysis was relieved obviously and bilirubin was reduced gradually after exchange transfusion. The infant was discharged in good health 13 days after birth with 135.0 g/L, 28.0 micromol/L and 10.7 micromol/L of Hb, total bilirubin and direct bilirubin, respectively. No sequelae were observed in a three-year follow-up. The result suggesting that the least incompatible blood is an alternative choice for exchange transfusion in severe HDN due to anti-Rh17 in case that Rh17 antigen-negative blood is unavailable.

Intraperitoneal transfusion for severe, early-onset rhesus disease requiring treatment before 20 weeks of gestation: A consecutive case series.

OBJECTIVE: To describe the management and perinatal outcomes of pregnancies affected by severe, early onset Rhesus isoimmunization treated with fetal intraperitoneal transfusions (IPTs). STUDY DESIGN: A ten-year consecutive case series of fetuses undergoing IPTs before 20 weeks gestation at Wessex Fetal Medicine Unit, Southampton, UK. Women with fetuses at risk of early onset fetal anaemia (before 20 weeks gestation) were identified from their obstetric history and maternal antibody levels at the time of booking. They were referred to our tertiary referral center. The decision to initiate transfusion was aided by middle cerebral artery peak systolic velocity as an indicator of fetal anaemia. No fetus was hydropic at the first transfusion. IPTs were commenced from as early as 15 weeks gestation in fetuses with difficult vascular access and performed regularly using this method until the cord could be successfully entered for intravascular transfusions. The main outcome measures were procedure and non procedure-related perinatal losses. RESULTS: 11 fetuses underwent 45 IPTs. 10/11 (91%) were delivered after 33 weeks gestation. There was one perinatal loss 1/11 (9%: 95% C.I 1.6-38%) from a cord accident during intravascular transfusion at 26 weeks gestation. There were no procedure related fetal losses or complications at the time of early IPTs. CONCLUSIONS: Previous studies report a perinatal loss rate with early intrauterine transfusion of 24% in gestations below 20 weeks using the intravascular route. This series suggests that intraperitoneal transfusion can be a safe and effective treatment for severe fetal anaemia at early gestations where vascular access is difficult.

Mirror syndrome due to anti-Jra alloimmunization.

OBJECTIVE: We report a case of fetal hydrops and mirror syndrome in a pregnancy with anti-Jra alloimmunization. CASE REPORT: A 34-year-old multiparous woman (G3P2) at 29 weeks of gestation had complications which included generalized edema and mild dyspnea. An indirect Coombs test was positive for anti-Jra antibodies. A blood examination showed hemodilution and elevated human chorionic gonadotropin. An ultrasound examination showed fetal hydrops with cardiomegaly and polyhydramnios. The patient delivered a pale and edematous infant by cesarean section and laboratory tests showed that the neonate had severe anemia (Hb 4.4 g/dL). A direct Coombs test was also positive. Microscopic examination of the placenta revealed diffuse villous edema. A genetic test for the ABCG2 gene showed the homozygous point mutation c.376C > T (376TT) in the mother, while her three offsprings all exhibited 376CT heterozygosity. CONCLUSION: The potential risk of severe fetal hydrops and mirror syndrome should be recognized in pregnancies with anti-Jra alloimmunization.

Etiology of nonimmune hydrops fetalis: a systematic review.

Hydrops fetalis (HF) indicates excessive fluid accumulation within the fetal extravascular compartments and body cavities. HF is not a diagnosis in itself but a symptom, and the end-stage of a wide variety of disorders. In the era before routine immunization of Rhesus (Rh) negative mothers, most cases of hydrops were due to erythroblastosis from Rh alloimmunization, but nowadays, nonimmune hydrops fetalis (NIHF) is more frequent, representing 76-87% of all described HF cases. We performed a systematic review of the pertinent literature based on the QUality Of Reporting Of Meta-analyses (QUOROM) recommendations, using a QUOROM flowchart and QUOROM checklist. At initial screening 33,345 articles were retrieved. The various inclusion and exclusion criteria aimed at obtaining data that were as unbiased yet as complete as possible decreased the numbers dramatically, and eventually a total of 225 relevant NIHF articles were identified, describing 6,361 individuals. We established 14 different diagnostic categories and provide the pathophysiologic background of each, if known. All 6,361 patients were subclassified into one of the following diagnostic categories: Cardiovascular (21.7%), hematologic (10.4%), chromosomal (13.4%), syndromic (4.4%), lymphatic dysplasia (5.7%), inborn errors of metabolism (1.1%), infections (6.7%), thoracic (6.0%), urinary tract malformations (2.3%), extra thoracic tumors (0.7%), TTTF-placental (5.6%), gastrointestinal (0.5%), miscellaneous (3.7%), and idiopathic (17.8%).

Maternal steroid therapy for fetuses with immune-mediated complete atrioventricular block: a systematic review and meta-analysis.

INTRODUCTION: To explore the effect of maternal fluorinated steroid therapy on fetuses affected by immune-mediated complete atrio-ventricular block (CAVB) in utero. MATERIAL AND METHODS: Pubmed, Embase, Cinahl, and ClinicalTrials.gov databases were searched. Only studies reporting the outcome of fetuses with immune CAVB diagnosed on prenatal ultrasound without any cardiac malformations and treated with fluorinated steroids compared to those not treated were included. The primary outcome observed was the regression of CAVB; secondary outcomes were need for pacemaker insertion, overall mortality, defined as the occurrence of either intrauterine (IUD) or neonatal (NND) death, IUD, NND, termination of pregnancy (TOP). Furthermore, we assessed the occurrence of all these outcomes in hydropic fetuses compared to those without hydrops at diagnosis. Meta-analyses of proportions using random effect model and meta-analyses using individual data random-effect logistic regression were used to combine data. RESULTS: Eight studies (162 fetuses) were included. The rate of regression was 3.0% (95%CI 0.2-9.1) in fetuses treated and 4.3% (95%CI 0.4-11.8) in those not treated, with no difference between the two groups (odds ratio (OR): 0.9, 95%CI 0.1-15.1). Pacemaker at birth was required in 71.5% (95%CI 56.0-84.7) of fetuses-treated and 57.8% (95%CI 40.3-74.3) of those not treated (OR: 9, 95%CI 0.4-3.4). There was no difference in the overall mortality rate (OR: 0.5, 95%CI 0.9-2.7) between the two groups; in hydropic fetuses, mortality occurred in 76.2% (95%CI 48.0-95.5) of the treated and in 23.8% (95%CI 1.2-62.3) of the untreated group, while in those without hydrops the corresponding figures were 8.9% (95%CI 2.0-20.3) and 12% (95%CI 8.7-42.2), respectively. Improvement or resolution of hydrops during pregnancy occurred in 76.2% (95%CI 48.0-95.5) of cases treated and in 23.3% (95%CI 1.2-62.3) of those nontreated with fluorinated steroids. CONCLUSIONS: The findings from this systematic review do not suggest a potential positive contribution of antenatal steroid therapy in improving the outcome of fetuses with immune CAVB.

Hydrops fetalis caused by homozygous alpha-thalassemia and Kidd antigen alloimmunization in a Chinese woman.

We report a case with hydrops fetalis resulting from homozygous alpha-thalassemia and alloimmune hemolytic anemia due to anti-JK3. This is one of the few cases with immune- and nonimmune-induced hydrops fetalis.

Successful management of a hydropic fetus with severe anemia and thrombocytopenia caused by anti-CD36 antibody.

Cases of CD36 deficiency are not rare in Asian populations, foetal and neonatal alloimmune thrombocytopenia (FNAIT) caused by anti-CD36 isoantibodies appears more frequent than other HPA alloantibodies. However, little is known about the treatment of anti-CD36 mediated FNAIT in this region. A Chinese male foetus, whose mother had a history of multiple intrauterine foetal demise and/or hydrops, was diagnosed with severe FNAIT at 27 weeks of gestational age. Immunological analysis revealed total absence of CD36 on platelets and monocytes from mother, caused by a 329-330delAC mutation of the CD36 gene. Anti-CD36 and anti-HLA class I antibodies were detected in the maternal serum, whereas only anti-CD36 isoantibodies were detectable in the foetal blood sample. Serial intrauterine transfusions with red blood cells (RBC) and platelets from a CD36null donor were performed to improve the severe anaemia and thrombocytopenia. The baby (2250 g; Apgar scores 10) was delivered vaginally at 32 weeks of gestation with normal haemoglobin (186 g/L) but low platelet count (48 × 109/L). After 2 days the platelet count rose to 121 × 109/L. This report suggests that intrauterine transfusions with compatible RBC and CD36null platelets are useful in preventing the deleterious clinical effects of anti-CD36-mediated severe FNAIT.

Two sibling cases of hydrops fetalis due to alloimmune anti-CD36 (Nak a) antibody.

Two female sibling cases, who were born to a CD36 deficient mother, were presented with Coombs' test-negative hydrops. The alloimmune anti-CD36 (Nak(a)) antibody was accidentally found in the mother's serum after an episode of anaphylactic shock with thrombocytopenia, which occurred in an individual receiving fresh frozen plasma prepared from the mother's donated blood. The mother was then diagnosed as having type II CD36 deficiency, lacking CD36 on both platelets and monocytes, while both of her daughters were CD36 positive. Analyses of the CD36 gene revealed that the mother was a compound heterozygote for the CD36 gene mutation with a novel C --> T transition at nt 1366 in exon 12, corresponding to Arg386Trp, and a known 12bp deletion at nt 1438-1449 in exon 13. On the other hand, both patients, who showed half the normal level of CD36 on platelets and monocytes, were heterozygote with one mutation at Arg386Trp. The anti-CD36 antibody in the mother seemed to be responsible for the hydrops fetalis observed in her daughters, because the IgG isolated from the mother's serum showed suppressive effects on the CFU-E colony formation of CD34+ cells from a control donor. This is the first case report of hydrops fetalis caused by an alloimmune anti-CD36 antibody.

Chicken egg yolk antibodies specific for the gamma chain of human hemoglobin for diagnosis of thalassemia.

Immunoglobulin Y (IgY) technology was used to generate anti-hemoglobin Bart's (Hb Bart's) IgY antibodies (Abs) for development into an enzyme-linked immunosorbent assay (ELISA) test for thalassemia diagnosis. Hb Bart's purified from the hemolysate of a patient with Hb Bart's hydrops fetalis (homozygous alpha-thalassemia) was used to immunize a chicken via the pectoralis muscle. After water dilution and sodium sulfate precipitation, 40 to 70 mg of IgY could be extracted from an egg. IgY, first detected in sera 2 weeks after immunization, reached the highest titer at week 4, and the titer remained stable for at least 2 weeks before declining. The pattern of Ab response in the yolk was the same as in the serum but was somewhat delayed. The IgY Abs produced reacted with gamma globin, Hb Bart's, Hb F, normal cord hemolysate (Hbs F plus A), and Hb Bart's hydrops fetalis (Hbs Bart's plus Portland) and to a lesser degree with beta globin, Hb A, Hb A2 and adult hemolysate (Hbs A plus A2), but the Abs did not react with alpha globin. Immunoaffinity purification with Hb A coupled to Sepharose was used to isolate an unbound IgY that reacted with Hb F, Hb Bart's, and gamma globin, and this IgY was used to develop an ELISA test for thalassemia diagnosis. The results of direct ELISA analyses of 336 hemolysate samples from individuals with various known thalassemia genotypes and phenotypes and from healthy individuals confirmed the specificity of the polyclonal Abs for Hbs containing Hb F and Hb Bart's. This specificity, which was due to the Abs' strong reactivity in cases of pathologic thalassemic diseases and weak reactivity in cases of nonpathologic thalassemic diseases, depended on the levels of Hb Bart's and Hb F.

Recurrent non-immune fetal hydrops: A case report.

INTRODUCTION: Recurrent non-immune fetal hydrops (NIH) has been reported in the literature but is a rare entity, with fewer than 6 reported cases so far. It has been postulated to be related to a recessive gene. CLINICAL PICTURE: We report a case of recurrent fetal hydrops in a multigravida with no medical history of note. She presented in her current pregnancy with a significant history of having 4 (out of 7) previous pregnancies affected by hydrops. TREATMENT: All the affected pregnancies resulted in mid-trimester pregnancy termination (MTPT) following diagnosis in the second trimester. Previous investigations for hydrops did not yield any obvious cause. OUTCOME: Her most recent pregnancy was unaffected. We discuss the possible differential diagnoses and the likelihood of autosomal recessive metabolic diseases being the aetiological factor. CONCLUSION: Rare causes of fetal hydrops need to be excluded in cases of recurrent non-immune hydrops with no obvious aetiology following routine investigations.

Management of severe fetal anemia by Doppler measurement of middle cerebral artery: are there other benefits than reducing invasive procedures?

OBJECTIVE: Doppler measurement of peak velocity of systolic blood flow in the middle cerebral artery (PVS-MCA) can safely replace invasive testing in the diagnosis of fetal anemia in Rh-alloimmunized pregnancies and PSV-MCA is now the reference technique. However, no study has evaluated its impact in antenatal care and in survival rate. Our objective was to evaluate the impact of the measurement of PVS-MCA in antenatal management and neonatal outcome in maternal red cell alloimmunization requiring in utero transfusion (IUT). STUDY DESIGN: Retrospective study between January 1999 and January 2013. We excluded all cases of hydrops without follow-up before first IUT. From 1999 to 2006, an IUT was indicated on the optical index at 450 nm (Period 1) and was then replaced by the use of PVS-MCA (Period 2). RESULTS: 77 patients were included, 39 in Period 1 (104 IUT) and 38 in Period 2 (89 IUT). 5 cases of hydrops fetalis (12.8%) were diagnosed during the follow up in Period 1 and none during Period 2. The average number of IUT, the delays between 2 IUT and between last IUET and birth were comparable. The total rate of complication per IUT during the first period was 9.6% vs 1.1% during the second one (p=0.01). The overall survival rate in our population was 34/39 (86.8%) during Period 1 vs 38/38 (100%) during Period 2. CONCLUSION: PSV-ACM allowed an improved monitoring with fewer occurrences of hydrops. Conversely, it did not modify antenatal management and timing of delivery.

Design and production of a target-specific monoclonal antibody to parvovirus B19 capsid proteins.

Native parvovirus B19 was used as antigen to produce a mouse monoclonal antibody, R92F6, which reacted with B19 VP1 and VP2, neutralised the virus in bone marrow culture, and labelled infected cells in paraffin-embedded tissues from cases of B19-related fetal hydrops. The B19 epitope recognised by R92F6 (amino acids 328-344 from the amino terminal region of B19 VP2) appears to be highly conserved, since these tissue specimens were obtained over a 13 year period from widely spaced locations in the UK. This epitope was synthesised as a peptide (S7b) which was used as antigen to produce a mouse monoclonal antibody, 3H8, which specifically reacted with the B19 capsid proteins VP1 and VP2 in immunofluorescence and immunoblot assays. 3H8 was also capable of labelling formalin-fixed, paraffin-embedded, B19-infected fetal tissue and was shown to be of the same isotype as R92F6 (IgG1). Highly conserved epitopes derived from conserved amino acid sequences are valuable in the diagnosis of infectious disease. If these can be recognised and accurately synthesised, the production of specific mouse monoclonal antibodies may be possible for many human pathogens. Considering the vast amount of sequence data available in the literature, this approach seems to be both feasible and of wide potential.

Nonimmune hydrops fetalis.

In summary, NIHF is a heterogenous disorder resulting from a vast number of underlying pathologies. A thorough evaluation should be performed in all cases to attempt to establish the etiology. This requires a systematic approach that should logically proceed from least to most invasive testing. Despite increasing availability of treatment for some causes of NIHF, the prognosis for this condition in general remains poor. In cases of fetal or neonatal demise, autopsy should be encouraged to aid in confirming or making a diagnosis. It is especially important to rule out potentially treatable conditions, as well as genetic disorders with a risk of recurrence in future pregnancies.

Intrauterine treatment on non-immune hydrops fetalis.

In 51 cases with non-immunologic hydrops fetalis (NIHF), perinatal management was performed based on our protocol. Twenty-two cases were treated by albumin and/or packed red cell (PRC) injection into the fetal abdominal cavity, and 9 cases by transplacental digitalization. Among the cases treated by albumin and/or PRC injection, 6 of 8 cases without pleural effusion recovered in utero, and all 6 cases are alive. However, of 14 cases with pleural effusion, none recovered in utero, and only one case is alive. Of 9 cases treated by transplacental digitalization, 2 cases recovered in utero, and only one case is alive. All fetuses with congenital heart anomaly died. This evidence indicates that albumin and/or PRC injection into the fetal abdominal cavity is an effective procedure for in utero treatment of NIHF without pleural effusion, but suggests that in NIHF resulting from either congenital heart anomaly and/or heart failure, the survival rate may not be increased by transplacental digitalization.

The role of the fetal immune system in the pathogenesis of RhD-hemolytic disease of newborns.

Data in the literature and author's research regarding the role of the immune reaction of fetuses and newborns in the pathogenesis of RhD conflict and hemolytic disease of newborns was analyzed. In this disease, the immune response of fetuses and newborns is shown to develop under the effects of maternal antigens, including RhD IgG, which cross the placenta. One of the results is the formation of immune complexes (ICs) between the maternal antigens and fetal IgM. In the intensive immune reaction, these ICs are removed from the infants at a high rate. As a result, the intensity of erythrocyte destruction, the degree of anemia and hyperbilirubinemia decrease. Various forms of HDN are of different intrauterine duration: from a few days in the icteric form without anemia to a month or more, in the hydropic form. In the latter form, decompensation of the immune system develops; extravascular erythroclasia by macrophages is replaced by intravascular lysis of erythrocytes. We suggest some methods to determine the fetal condition and a cure for the most severe cases of HDN, as well as a way of decreasing RhD-sensitization in women. These suggestions may be of interest to specialists in pediatrics and obstetrics and may be of clinical use.

Haemolytic disease of the newborn due to anti-K antibodies.

We describe a case of intrauterine foetal death at 32 weeks gestation with signs of hydrops foetalis without erythroblastosis. The hydrops foetalis appeared to be caused by blood group incompatibility. Irregular antibodies of anti-K specificity were found in the serum of the mother, while the father was positive for the K-antigen. The antibody dependent cell-mediated cytotoxicity (ADCC) test with serum of the mother was positive. This case shows the characteristics of haemolytic disease of the newborn by anti-K. Moreover, it underlines the necessity of both adequate follow-up of individual cases, and screening for irregular antibodies in all pregnancies as well as central registration of screening results.