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1 PERICONCEPTIONAL RISK FACTORS OF SPINA BIFIDA AMONG EGYPTIAN 2 POPULATION: A CASE-CONTROL STUDY 3 4 Moutaz Elsherbini, MD(1)*. ([email protected]) 5 Wafaa Ramadan, MD(1). ([email protected]) 6 Rasha Elkomy,MD(1). ([email protected]) 7 Omneya Helal,MD(2)*. ([email protected]) 8 Dina Latif Hatem,MD(1).([email protected]) 9 Hassan Gaafar, MD(2). ([email protected]) 10 11 12 13 14 (1) Lecturer of Obstetrics and Gynecology, Kasr Alaini, Faculty of Medicine, Cairo University, Egypt (2) Assistant professor of Obstetrics and Gynecology, Kasr Alaini, Faculty of Medicine, Cairo University, Egypt 15 16 17 18 Corresponding Author: 19 Hassan Mostafa Gaafar 20 4-Gamal salem St ,Dokki, Cairo 21 Tel: 01224022330 22 23 E-mail: [email protected] 24 25 26 27 28 29 1 30 Periconceptional risk factors of spina bifida among Egyptian population: 31 a case-control study 32 Abstract 33 Objective: To study the preconceptual & early conceptional risk factors that may 34 predispose to the development of spina bifida (SB) malformation among Egyptian 35 population. 36 Methodology: The study involved 197 pregnant women undergoing fetal anatomy 37 scan; 97 women proved to have fetal SB and 100 women with normal fetuses as a 38 control group. The control group was recruited randomly in the same period from 39 patients undergoing anatomical scan. Risk factors that might lead to SB were 40 investigated including maternal age, gravidity, parity, residence, history of diabetes 41 mellitus or drug intake, smoking, infections, exposure to X-ray, history of congenital 42 anomalies in other offspring, parental consanguinity, positive family history, folate 43 supplementations. 44 Results: SB affected the lumbo-sacral region in the majority of cases (89.7%). It was 45 associated with hydrocephalus in 66 cases (68%), polyhydramnios in 12 cases (12.4%). 46 The SB group showed significantly higher parity (p = 0.005), more frequent history of 47 drug intake (p < 0.001), higher frequency of infection with CMV (p = 0.004) and HSV (p 48 = 0.013) and less proportion of folate supplementation (p < 0.001). 49 Conclusion: The rate of spina bifida in the tested group was 5 per 1000. Risk factors 50 were lack of folate supplementation & history of antiepileptic drugs intake. This 51 association raises the hypothesis that the disturbed folate/homocysteine metabolism 52 is the underlying mechanism for SB development. 53 54 Keywords: Spina bifida, 3D ultrasound, Hydrocephalus, Consanguinity 2 55 Introduction: 56 Spina bifida (SB) is a serious neural tube birth defect that approximately 57 affects 35/100000 live births 58 dysraphisms as it represents about 90% of all spinal defects 59 from failure of proper fusion in the spinal regions of the neural tube 60 during early embryogenesis (within the first 28 days following conception) 61 (3) 62 Neural tube defect (NTD) incidence varies significantly across time and 63 among different geographic areas within the same time with the highest 64 incidences among Irish, Indian, Mexican and Northern China populations 65 (3-5) 66 however it might decrease due to maternal folic acid supplementation 67 together with the prenatal diagnosis and termination of affected 68 pregnancies (7,8). 69 Babies with SB are compatible with life but handicapped with increased 70 morbidity and mortality risks throughout their lives. They usually suffer 71 from mental retardation, motor and sensory dysfunction of lower limbs 72 and inability to control the anal and urethral sphincters (9). 73 Many surgical procedures are usually required to close the vertebrae, 74 stabilize the joints and drain the associated hydrocephalus but without 75 complete restoration of normal life and consequently, when parents faced 76 with diagnosis of SB prenatally, nearly ¾ of them decide to induce 77 abortion (10). 78 Like other NTD, SB appears to have a multifactorial etiology, with the 79 environmental factors, lifestyle; social and cultural habits interacting with (1) . It is the most common form of spinal (2) . SB results . . In Europe and North America, SB birth prevalence is 0.3-0.6/1000 3 (6) , (11) 80 the genetic susceptibilities to determine the risk of development . 81 Particularly, maternal nutrition has been recognized as an important 82 predisposing factor 83 recommended daily dose of 400 μg, preconceptually and in the first 84 trimester was found to reduce the risk of NTD development 85 consequently, US and Canada in 1998 started folic acid fortification of 86 enriched cereal grains (13). 87 The risk increases to 3% with one previous NTD-affected pregnancy and to 88 10% with two previous NTD-affected pregnancies. Other risk factors 89 include the maternal use of antiepileptic drugs (e.g., valproic acid) or folic 90 acid antagonists (e.g., methotrexate), preconceptual maternal diabetes 91 and obesity and maternal exposure to high temperature in early 92 pregnancy (e.g., fever, hot tubs and saunas) 93 recognizable risk factors in 90% of cases (16). 94 NTDs are the easiest congenital anomalies to diagnose prenatally 95 Traditionally, elevated maternal serum α-fetoprotein (α-FP) followed by 96 sonographic scanning were the diagnostic tools (18). More recently, Routine 97 antenatal fetal anatomy scan using 2D & 3D is an efficient screening tool 98 of SB (detection rate up to 95-100%) (19). 99 The aim of this work is to study the preconceptual and early conceptional 100 risk factors that may predispose to the development of SB malformation 101 among Egyptian population. (11) . Maternal folic acid supplementation, with 102 103 4 (12) and (14,15) . However, there are no (17) . 104 Subjects and Methods: 105 In the current study, 18653 pregnant women were scanned in the 106 ultrasound department in Kasr-El-Aini University hospital (Cairo University, 107 Egypt) from the period of June 2011 to June 2014 after being approved by 108 the hospital ethics research committee. All of them were referred from the 109 obstetric outpatient clinic for routine obstetric ultrasound or for suspected 110 anomalies recorded at any trimester of pregnancy. 111 Ultrasound was done by two expert operators with the use of high 112 resolution ultrasound unit with 3-5 MHz transabdominal transducers 113 (General electric voluson Pro-V 730). Every ultrasound scan took about 15- 114 20 minutes with detailed anatomical scanning of the fetus according to the 115 ISUOG guidelines (20). 116 Cases of spina bifida (97 patients) were identified and complementary 3D 117 ultrasound was done by senior operator with documentation of data as 118 regards 1) the presence or absence of hydrocephalus 2) Level of the lesion 119 3) proper assessment of the liquor volume. Control patients (100 patients) 120 were enrolled randomly in the same period in which the fetal anatomy 121 scan proved the presence of non-malformed fetus. (fig 1-4) 122 Exclusion criteria include maternal age > 42 or <18 years old, presence of 123 multiple fetal anomalies, non-viable fetus, multiple gestation. Risk factors 124 were identified by simple verbal questions as regard: maternal age, 125 gravidity, parity, residence (urban or rural), history of congenital 126 anomalies in other offspring, parental consanguinity, positive family 127 history, 128 supplementations. Other risk factors assessed include preconceptual or maternal history of diabetes 5 mellitus and folic acid 129 first trimester history of antiepileptic drugs (AEDs) intake, smoking, 130 exposure to X-ray and infections namely; Rubella, CMV and HSV, which is 131 confirmed by presence of virus- specific clinical presentation and elevated 132 serum IgM (1ry infection) or the rising titer of virus- specific serum IgG 133 (recurrent infection). 134 135 6 136 Results: 137 SB affected the lumbo-sacral region in most of the studied cases; 87 out of 138 the 97 fetuses (89.7%). Only 10 cases (10.3%) had thoracic SB. It was 139 associated with hydrocephalus in 66 cases (68%). Polyhydramnios was 140 significantly more common in SB cases (p = 0.013); it was found in 12 cases 141 (12.4%) compared to 3 cases (3%) of the control group. 142 Table 1 shows the studied risk factors of SB in the two groups. The SB 143 group showed significantly higher parity (p = 0.005), more frequent history 144 of AEDs intake (p < 0.001), higher frequency of infection with CMV (p = 145 0.004) and Herpes virus (p = 0.013) and less proportion of folate 146 supplementation (p < 0.001). 147 148 7 149 Discussion: 150 This study demonstrated a rate of diagnosis of spina bifida (SB) of 5 per 151 1000 examined pregnant women; mainly affecting the lumbo-sacral region 152 (89.7%). SB was significantly associated with higher parity (p = 0.005) & 153 more frequent history of AEDs intake (p < 0.001), CMV infection (p = 154 0.004) and Herpes virus (p = 0.013). Also, significantly lower proportion of 155 those with SB had folate supplementation during the current pregnancy (p 156 < 0.001). 157 158 We cannot rely on the proportion of SB found in this study as an indicator 159 of the incidence of the defect among Egyptian population. This study is a 160 hospital based one performed in a tertiary referral center. The exact 161 incidence in Egypt needs a population based study which is lacking so far. 162 The prevalence of SB is characterized by geographic and temporal 163 variation in addition to racial and ethnic differences within location and 164 time (21). The average worldwide incidence of SB is 1 case per 1000 births. 165 Prevalence of neural tube defects (NTDs) ranges from 0.7 in central France 166 to 7.7 in the United Arab Emirates and 11.7 in South America per 10,000 167 births. In the United States, SB incidence is 7 per 10,000 live births (22). 168 However, the birth prevalence of NTDs is influenced by the accessibility of 169 prenatal diagnostic tools and the use of elective pregnancy termination 170 (23) 171 The etiology in most cases of SB is multifactorial, involving genetic, 172 environmental and dietary factors. In the current study, lack of folic acid 173 supplementation was associated with higher frequency of SB (p < 0.001). . 8 174 Similarly, many studies have reported significant relation between the low 175 consumption of folic acid and NTDs in Europe (24) and Asia (25). It is believed 176 that nearly half of cases of NTDs are related to a nutritional deficiency of 177 folic acid, but the underlying mechanism is not clear. A genetic or 178 nutritional defect in homocysteine metabolism was suggested 179 research articles addressed the role of correction of folic acid deficiency 180 for prevention of primary and recurrent NTDs (27). However, the underlying 181 protective mechanism is still not identified, but possibly through their 182 effects on genes that regulate folate transport and metabolism (28). 183 According to the CDC, mandatory fortification of enriched cereal grain 184 products with folic acid in the USA in 1998 resulted in 22.9% reduction of 185 the yearly incidence of SB 186 supplementation between 205 mothers of SB cases and 6357 mothers of 187 controls; authors estimated a 13% reduced odds of SB for each 100 μg 188 increase in daily folate consumed 189 European health authorities recommend folic acid supplementation of 400 190 μg for pregnant women to prevent NTDs 191 studies involving 6105 women confirmed the protective effect of daily 192 folic acid supplementation in prevention of NTDs (32). 193 In the current study SB was significantly associated with higher parity. 194 Relation between NTDs risk & parity is either U-shaped pattern (the risk is 195 higher for the lowest and highest number of births 196 proportionate (increased risk with increased parity) (34). 197 In the current study, history of antiepileptic drugs intake (AEDs), namely 198 Depakine (valproic acid sodium), Lamotrine (lamotrigine) and Tegretol (26) . Many (29) . In a large study, comparing folic acid (30) . Correspondingly, Most of the 9 (31) . A systematic review of 5 (33) or directly 199 (carbamazepine), was significantly more common in the SB group; 21.6% 200 vs. 1% in the control group (p < 0.001). These drugs are believed to be 201 associated with SB being anti-folate. 202 The association between SB and exposure to valproic acid during 203 pregnancy was reported in a retrospective study in France 204 subsequently confirmed in several studies 205 presented as the antiepileptic drug of choice during pregnancy 206 However, in utero exposure to Carbamazepine has been associated with 207 an increased risk of NTDs (39), but a recent systematic review showed that 208 the risk of SB development is significantly lower when compared to 209 valproic acid 210 its teratogenic effect in the form of dose-dependent growth retardation 211 and neurodevelopmental toxicity. SB and anencephaly were associated 212 with higher doses of the drug (41). 213 Lamotrigine is an anticonvulsant & mood stabilizer drug that has weak 214 antifolate properties 215 many researchers considered its use relatively safe as the rates of 216 congenital malformation in lamotrigine-exposed fetuses similar to those 217 observed in the general population 218 (45) 219 triplets on Lamotrigine treatment; however, they did not observe 220 significant changes in serum folate levels. Based on the above, 221 preconceptual & 1st trimesteric folic acid supplementation (in a daily dose 222 of 5 mg) had previously been recommended for all women taking AEDs (46). 223 But recently many studies have revealed that folic acid supplementation in (35) . This was (36,37) . Carbamazepine has been (38) . (40) . A recent animal study found that carbamazepine exerts (42) . It is rated Pregnancy Category Risk C, however, (43,44) . On the contrary, Candito et al. presented a case-report of double fetal NTDs in a patient pregnant with 10 224 this population group had not added any protective value (47), however, It 225 is still generally recommended that all women on AEDs should receive folic 226 acid supplementation (at a dose of at least 0.4 mg daily)(48). 227 Congenital cytomegalovirus (CMV) infection is the most prevalent viral 228 cause of congenital neurological disabilities in children that occurs in 0.6– 229 0.7% of all newborns worldwide (49). Vertical transmission can follow either 230 a primary or recurrent maternal infection with 90% of affected fetuses is 231 asymptomatic 232 primary infection & in symptomatic fetuses. Hearing loss, visual 233 impairment, or diminished mental and motor capabilities are the most 234 common sequelae 235 significantly more common in the SB group; 14.4% vs. 3% in the control 236 group (p < 0.001). Cannon MJ et al. reported that CMV infection is more 237 likely to occur among population of lower socioeconomic status 238 study was carried out at a University Hospital in which most of the 239 examined women are of low socioeconomic status. 240 This study has identified some maternal periconceptional risk factors 241 associated with an increased SB risk among Egyptian women. AEDs 242 administration was associated with increased risk while folic acid 243 supplementation was associated with reduced risk. This association raises 244 the hypothesis that the disturbed folate/homocysteine metabolism is the 245 underlying mechanism for SB development. Consequencely 246 information can be included as a part of pre-conception counseling. (50) . There is a higher risk for neurological damage after (50) . In the current study history of CMV infection was 247 248 11 (51) & our this 249 Conflict of Interest: 250 The authors have no conflicts of interest. 251 Authors’ contributions: 252 M.Elsherbini: Drafting the manuscript. 253 W.Ramadan: Ultrasound scanning and case choice. 254 R. Elkomy: Made contributions to conception and design. 255 O.Helal: Analysis and interpretation of data. 256 D. Hatem: Risk factors assessment. 257 H.Gaafar: Ultrasound scanning and case choice. 258 259 Acknowledgements: 260 The authors do not have anyone to acknowledge. 261 262 263 264 265 266 267 268 269 270 271 272 273 12 274 References: 275 1. Parker SE, Mai CT, Canfield MA, Rickard R, Wang Y, Meyer RE, Anderson P, Mason CA, Collins JS, Kirby RS & Correa A: Updated national birth prevalence estimates for selected birth defects in the United States, 2004–2006. Birth Defects Res. A: Clin. Mol. Teratol. 2010; 88, 1008-1016. 2. 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Rev Med Virol. 2010; 20:202-213. 419 420 421 422 16 423 Legends 424 425 Figure 1 426 2D ultrasound image of a sacral spina bifida 427 428 Figure 2 429 3D ultrasound image of spina bifida (same case) 430 431 Figure 3 432 Neonate with SB (same case) 433 434 435 436 437 438 439 440 441 442 443 444 445 446 447 17 448 Table 1: Maternal characteristics and risk factors of SB in the two studied 449 groups Normal Group (n = 100) 25.1±4.7 76.2±5.2 2 (0-5) 1 (0-4) p value Age (years) Maternal Weight (kg) Gravidity Parity Residence Rural Urban Positive Consanguinity Previous anomalies Family history of anomalies Diabetes Mellitus SB Group (n = 97) 26.6±6.5 77.0±6.0 2 (0-8) 1 (0-8) 55 (56.7%) 42 (43.3%) 48 (49.5%) 14 (14.4%) 26 (26.8%) 3 (3.1%) 46 (46.0%) 54 (54.0%) 39 (39.0%) 24 (24.0%) 19 (19.0%) 6 (6.0%) 0.133 X-ray exposure History of AEDs intake Smoking 9 (9.3%) 21 (21.6%) 8 (8.2%) 6 (6.0%) 1 (1.0%) 5 (5.0%) 0.386 < 0.001* 0.359 CMV Rubella Herpes virus 14 (14.4%) 7 (7.2%) 6 (6.2%) 3 (3.0%) 4 (4.0%) 0 (0.0%) 0.004* 0.326 0.013* Folate Supplementation 84 (86.6%) 100 (100.0%) < 0.001* 450 *Statistically significant 451 SB= Spina Bifida 452 CMV= Cytomegalovirus 453 18 0.052 0.301 0.758 0.005* 0.138 0.089 0.192 0.498