Background: Ghrelin is a pleiotropic hormone that regulates feeding and energy balance and stimulates growth hormone release. Ghrelin also exerts developmental and organizational effects during prenatal life. Objectives: The aim of this study was to determine ghrelin levels in cord blood of small for gestational age (SGA) infants and its association with GH (growth hormone) and IGF-1 levels (insulin-like growth factor-1). Methods: Cord blood sample was obtained from 31 SGA and 25 appropriate for gestational age (AGA) infants. Acylated ghrelin, GH, and IGF-1 levels were measured by enzyme-linked immunosorbent assay. Results: No significant differences were observed in ghrelin and GH concentrations between SGA and AGA infants. However, IGF-1 levels were significantly lower in SGA infants. Cord blood ghrelin was negatively correlated with the infants' birth weight (r = -0.33, P = 0.013); on the other hand, IGF-1 level was positively correlated with birth weight (r = 0.43, P = 0.002). Conclusions: IGF-1 has the most significant effect on intrauterine growth. Acylated ghrelin is detectable in cord blood and correlated with birth weight, suggesting a role in intrauterine development, but its level is not affected by intrauterine growth retardation. Â© 2016, Iranian Society of Pediatrics

Arab, Z.

Bordbar, A.

Choobdar, F.A.

Khalesi, N.

Khosravi, N.

Larijani, B.

Nourbakhsh, M.

English

eprints Iran University of Medical Sciences

J Compr Ped. 2016 November; 7(4):e42809.Published online 2016 November 12.doi: 10.17795/compreped-42809.Research ArticleAcylated Ghrelin, Growth Hormone and IGF-1 Levels in the Cord Bloodof Small for Gestational Age NewbornsNastaran Khosravi,1 Farhad Abolhasan Choobdar,1 Nasrin Khalesi,1 Arash Bordbar,1 Bagher Larijani,2Zahra Arab,3 and Mitra Nourbakhsh3,4,*1H. Aliasghar Hospital, Iran University of Medical Sciences, Tehran, IR Iran2Endocrinology and Metabolism Research Center, Endocrinology and Metabolism Research Institute, Tehran University of Medical Sciences, Tehran, IR Iran3Metabolic Disorders Research Center, Endocrinology and Metabolism Molecular-Cellular Sciences Institute, Endocrinology and Metabolism Research Institute, TehranUniversity of Medical Sciences, Tehran, IR Iran4Department of Biochemistry, School of Medicine, Iran University of Medical Sciences, Tehran, IR Iran*Corresponding author: Mitra Nourbakhsh, M.Sc., Ph.D, Department of Biochemistry, School of medicine, Iran University of Medical Sciences, 1417613151, Tehran, IR Iran. Tel/Fax:+98-21 88622742, +98 9122874740, E-mail: nourbakhsh.m@iums.ac.irmitra.nourbakhsh@gmail.comReceived 2016 October 05; Accepted 2016 October 09.AbstractBackground: Ghrelin is a pleiotropic hormone that regulates feeding and energy balance and stimulates growth hormone release.Ghrelin also exerts developmental and organizational effects during prenatal life.Objectives: The aim of this study was to determine ghrelin levels in cord blood of small for gestational age (SGA) infants and itsassociation with GH (growth hormone) and IGF-1 levels (insulin-like growth factor-1).Methods: Cord blood sample was obtained from 31 SGA and 25 appropriate for gestational age (AGA) infants. Acylated ghrelin, GH,and IGF-1 levels were measured by enzyme-linked immunosorbent assay.Results: No significant differences were observed in ghrelin and GH concentrations between SGA and AGA infants. However, IGF-1levels were significantly lower in SGA infants. Cord blood ghrelin was negatively correlated with the infants’ birth weight (r = -0.33,P = 0.013); on the other hand, IGF-1 level was positively correlated with birth weight (r = 0.43, P = 0.002).Conclusions: IGF-1 has the most significant effect on intrauterine growth. Acylated ghrelin is detectable in cord blood and corre-lated with birth weight, suggesting a role in intrauterine development, but its level is not affected by intrauterine growth retarda-tion.Keywords: Ghrelin, Growth Hormone (GH), IGF-1, Small for Gestational Age (SGA), Appropriate for Gestational Age (AGA),Intrauterine Growth1. BackgroundGhrelin is a 28-amino-acid peptide that is primarily se-creted by gastric mucosal cells and plays a key role in ap-petite regulation and energy balance. It was discoveredthrough its effect on growth hormone (GH) secretion, andthus was presumed to be effective in growth (1-3). Severalother activities have also been attributed to ghrelin (4, 5).In addition to its effects on GH secretion, ghrelin has alsobeen shown to stimulate appetite and induce adiposity (6).Small for gestational age (SGA) newborn infantsshowed reduced growth during the intrauterine period,which might have occurred because of fetal, maternal orenvironmental factors. These infants have low levels ofGH after birth in response to nutritional deficiency beforebirth (3). Although most of these infants may catch upgrowth rapidly after birth, various patterns of weight gainis observed in infancy, which may be related to feedingand fasting (2, 4). There is a strong association betweenvery low birth weight and infant mortality, which may bedue to extreme fetal growth retardation (7).Considering the important relationship between nu-trition and growth and the orexigenic function of ghrelin,this hormone seems to be closely associated with postna-tal growth. Presence of ghrelin in umbilical cord has beendemonstrated, but few studies have measured ghrelin lev-els in the umbilical cord of newborn infants (8, 9).With the assumption that umbilical cord ghrelin level,as well as GH and IGF-1 levels can indicate their levels dur-ing intrauterine period, the aim of this study was to deter-mine whether cord blood ghrelin, GH and IGF-1 levels aredifferent in SGA newborn infants compared to appropriatefor gestational age (AGA) newborn infants.2. Methods2.1. ParticipantsIn this cross-sectional study, 56 newborn infants bornat Akbarabadi hospital in Tehran were included. InfantsCopyright © 2016, Iranian Society of Pediatrics. This is an open-access article distributed under the terms of the Creative Commons Attribution-NonCommercial 4.0International License (http://creativecommons.org/licenses/by-nc/4.0/) which permits copy and redistribute the material just in noncommercial usages, provided theoriginal work is properly cited.Khosravi N et al.whose birth weight was below the 10th percentile for gesta-tional age were defined as SGA, and those with birth weightbetween 10th and 90th percentile were defined as AGA. Ac-cording to this definition, the infants were divided intotwo groups: 31 infants with SGA, and 25 with AGA. Bothgroups were included consecutively and by conveniencesampling.All the studied infants were devoid of any congenitalabnormalities, and had no history of maternal chronic dis-eases such as diabetes, autoimmune diseases, pregnancy-induced hypertension and pre-eclampsia. Newborns werefrom third gestation or less, and the mothers did notsmoke nor had any kinds of addiction.Demographic data, underlying conditions and 5thminute Apgar score were recorded. The height and weightof each infant were accurately measured and recorded inthe checklist.The Ethics Committee of Tehran University of MedicalSciences approved the study, and written informed con-sent was obtained from all mothers before cord blood sam-pling. All the procedures were in accordance with the eth-ical standards of the responsible committee on human ex-perimentation (institutional and national) and with theHelsinki Declaration of 1975, as revised in 2000.2.2. Biochemical AssaysVenous cord blood was collected immediately afterclamping the umbilical cord in EDTA tubes and kept at 4°Cbefore centrifugation. For ghrelin determination, PMSFwas added to the final concentration of 0.1 mg/mL as aprotease inhibitor. After centrifugation at 4°C, the sepa-rated plasma was acidified with HCl to a final concentra-tion of 0.05 N. All the samples were stored at 80°C untillater analysis. Acylated ghrelin was measured, using a com-mercial ELISA kit (DRG, USA), with intra- and inter-assay CVof 6.3% and 5.4%, respectively. Serum GH and IGF-1 weremeasured by chemiluminescence sandwich assay, usingdirectly coated magnetic microplates (Diasorin Ltd, UK).2.3. Statistical AnalysisThe differences in variables between groups were com-pared, using student’s t test. The chi-square test was usedto compare nominal variables between groups, and stu-dent’s t test was used for numeric variables. The relation-ship between different parameters was determined, usingPearson’s correlation. All the statistical analyses were per-formed, using SPSS Version 16 (SPSS Inc., Chicago, IL, USA). Pvalue < 0.05 was considered statistically significant.3. ResultsThe anthropometric and biochemical characteristicsof the neonates and their mothers are presented in Table 1.Overall, cord plasma ghrelin concentrations were 104.6±87 pg/mL (Range 12.58 - 508.54 pg/mL). As demonstrated inTable 1, although there was a trend towards higher ghrelinconcentration in SGA infants, no significant difference wasfound between the two groups in ghrelin concentration.Similarly, GH concentration was not statistically differentbetween SGA and AGA infants. IGF-1 level was significantlylower in SGA infants compared to AGA group. No signifi-cant differences were observed between male and femaleinfants in ghrelin, GH, and IGF-1 levels (Table 2).Cord blood ghrelin concentration was negatively cor-related with the infants’ birth weight (r = -0.33, P = 0.013).On the other hand, IGF-1 showed a significant positive cor-relation with birth weight (r = 0.43, P = 0.002). Never-theless, no relationship was found between GH and birthweight (P = 0.4). The relationship between GH, IGF-1, andacylated ghrelin was investigated and found to be non-significant. Mothers’ weights and their correlations withGH, IGF-1, and ghrelin were also investigated, but no signif-icant correlations were observed between them.4. DiscussionThe results of this study revealed that acylated ghre-lin is correlated with birth weight, and it is slightly butnot significantly higher in SGA infants. Among the stud-ied parameters, which are all effective in growth, IGF-1 ex-erts the most dramatic influence on growth consideringits low concentration in SGA infants, and its strong corre-lation with birth weight.Ghrelin has previously been shown to be elevated inanorexia and children with poor appetite (10) and dimin-ished in obesity, playing a role in the regulation of energyhomeostasis; thus, it is suggested that ghrelin representsthe chronic nutritional status (11, 12). The cord blood ghre-lin concentrations in SGA infants, compared to healthynewborns, have been reported in a few studies (8, 12, 13).All these studies reported that cord blood ghrelin con-centrations of SGA infants are greater than those of AGA in-fants, and it was suggested that ghrelin is affected by nu-tritional status during the fetal life. However, these studieshave investigated total ghrelin levels despite the fact thatacylated form of the ghrelin is the active form of the hor-mone, and acylation is mandatory for the biologic effectsof ghrelin on GH secretion and for binding to the recep-tor (14). Umbilical cord acylated ghrelin concentration islower than maternal blood, and the placenta plays a rolein the process of acylation (15). Thus, it is plausible that2 J Compr Ped. 2016; 7(4):e42809.Khosravi N et al.Table 1. Characteristics of the Neonates and Their MothersaSGA (31) AGA (25) Total (56) P ValueSex (M/F) 15/16 7/18 22/34 0.1Gestational age, w 35.5± 3 37.9± 2.5 36.6± 3 0.002bBirth weight, g 2177.6± 703.2 3247.2± 575.2 2672.7± 837.2 < 0.0001bGravid 1.9± 1.3 2.3± 1.1 2± 1.2 0.2Maternal age, y 27.1± 7 29.6± 5.9 28.2± 6.6 0.1APGAR 5 9.6± 0.6 9.9± 0.3 9.7± 0.5 0.06Ghrelin, pg/mL 121.26± 109.4 83.9± 40.2 104.6± 87 0.08GH, ng/mL 49.6± 40.3 54.7± 24.9 51.8± 34.2 0.58IGF-1, ng/mL 32.2± 32 59.3± 35.9 44± 36 0.008bAbbreviations: AGA, appropriate for gestational age; GH, growth hormone; IGF-1, insulin-like growth factor-1; SGA, small for gestational age.aValues are expressed as mean± SD.bSignificant difference.Table 2. Comparison of the Levels of ghrelin, GH and IGF-1 between Female and MaleInfantsFemale (34) Male (22) P ValueGhrelin, pg/mL 109.9± 94.9 96.3± 74.7 0.57GH, ng/mL 56.2± 32.6 45.2± 36.4 0.27IGF-1, ng/mL 49.3± 36.9 35.9± 33.9 0.2the acylated ghrelin changes may be different from thoseof total ghrelin. Iniguez et al. investigated the serum lev-els of ghrelin in SGA and AGA infants after fasting and in-travenous injection of glucose and found that after fasting,serum levels of ghrelin were not different between the SGAand AGA groups (16).IGF-1 is one of the main factors responsible for regu-lating the fetal growth (17). In this study, IGF-1 levels werefound to be significantly lower in SGA infants comparedto AGA newborns. Consistent with our findings, Orbaket al. have also demonstrated that IGF-1 levels are signifi-cantly diminished in SGA neonates (18). This study revealeda significant positive correlation between IGF-1 and birthweight in contrast to ghrelin, which was negatively corre-lated with birth weight. The levels of both of these hor-mones seem to be altered in response to nutritional sta-tus. Increased ghrelin level in anorexia is suggested to bea compensatory response to undernutrition (19). On theother hand, it has been shown that intake of proteins thatare rich in essential amino acids is positively associatedwith serum IGF-1 (20) and although total calorie restrictiondoes not affect IGF-1 levels, protein intake is a key determi-nant of circulating IGF-1 levels in humans (21).Our study found that ghrelin, GH, and IGF-1 concentra-tions were similar in male and female infants. In SGA in-fants, these hormones were not different in the two gen-ders, but in AGA group, GH of female infants was signif-icantly higher than that of the male infants. Consistentwith our findings, Chanoine et al. found no significant dif-ference between male and female infants (22).In conclusion, these results suggest that minimal in-crease in ghrelin levels could be the consequence ofgrowth restriction, but it may be caused by significant re-duction in IGF-1. Thus, IGF-1 may be a suitable target formanaging intrauterine growth.AcknowledgmentsThis study was financially supported by a grant (num-ber: 1392-01-104-1617) from Endocrinology and MetabolismResearch Institute, Tehran University of Medical Sciences.FootnoteConflict of Interest: The authors declare no conflict of in-terest.References1. Cunha SR, Mayo KE. Ghrelin and growth hormone (GH) secreta-gogues potentiate GH-releasing hormone (GHRH)-induced cyclicadenosine 3’,5’-monophosphate production in cells expressingtransfected GHRH and GH secretagogue receptors. Endocrinol-ogy. 2002;143(12):4570–82. doi: 10.1210/en.2002-220670. [PubMed:12446584].2. 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Acylated ghrelin, growth hormone and IGF-1 levels in the cord blood of small for gestational age newborns

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Acylated ghrelin, growth hormone and IGF-1 levels in the cord blood of small for gestational age newborns

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