Relation of Vascular Endothelial Growth Relation of Vascular Endothelial Growth Factor and Insulin Like Growth factor-1 to Factor and Insulin Like Growth factor-1 to

the Development of Retinopathy in the Development of Retinopathy in Premature InfantsPremature Infants

Prof. Dr: Enas Raafat

Prof of Child Health

National Research Center

IntroductionIntroduction

Retinopathy of prematurity (ROP) has been identified, as a retinal vasoproliferative condition that evolves into five stages.It is initiated by retinal vascular growth retardation.

Retinopathy of prematurity is one of the important causes of childhood blindness worldwide.

The blood vessels in the retina of premature and/or low birth weight infants are immature and underdeveloped.

Following delivery, the blood vessels continue to grow and spread throughout the retina. These abnormal blood vessels are fragile and can leak, scarring the retina and pulling it out of position leading to retinal detachment.

IntroductionIntroduction

IntroductionIntroductionThe abnormal vascular changes are divided into five stages:

Stage 1 ROP with demarcation line

Stage 2 ROP consisting of a ridge

Stage 3 :consisting of a ridge with extra retinal fibrovascular proliferation

Stage 4,5: refers to a partial and complete retinal detachment

IntroductionIntroduction

IntroductionIntroduction

Various risk factors to which the neonates are exposed, i.e. oxygen therapy, apnea, anemia,

etc… may aggravate the insult to which the under developed or premature eye is exposed to.

It has been proposed that vasculogenesis is the result of complex interactions between growth factors (cytokines like IGF-1 and VEGF) produced both locally and systemically.

IntroductionIntroduction

Insuline like growth factor -1 ( IGF-1) is important for normal retinal vascular development, its deficiency is associated with lack of vascular growth, leading for subsequent proliferative ROP.

Vascular endothelial growth factor( VEGF ) act as survival factor for newly formed capillaries in the developing retina and suggest an important role in the pathogenesis of human ROP.

Sequence of ROP StagesSequence of ROP Stages

Aim of the StudyAim of the Study

We aimed to evaluate the role of cytokine levels (IGF-1 and VEGF) in serum of premature infants as possible diagnostic markers for ROP.

We also aimed to analyze other factors which may act as risk factors for occurrence and severity of ROP, like birth weight and gestational age.

Patients and MethodsPatients and Methods

During the period from May 2008 to August 2009, 83 premature neonates who were admitted to the NICU of Almaza charity neonatal care unit in Cairo were included in this study.

This work was done in corporation between, Child Health and Clinical Pathology departments (NRC) and Ophthalmology department, Cairo University.

Patients and MethodsPatients and Methods

Inclusion Criteria:

All neonates included in the study were prematures. They were classified into two groups:

– Group A (cases): Premature neonates developing ROP (n = 34), they were subdivided to:

1- Mild cases (those with stages I and II ROP) 2-Severe cases (those with stages III and V ROP)

– Group B (controls): Premature neonates without ROP (n = 49)

Patients and MethodsPatients and MethodsExclusion Criteria:

– Fullterms.– Neonates with obvious congenital malformations.– Neonates with other causes for retinopathy.

All babies were examined during their stay in the NICU by the attending staff and also ophthalmic examination was performed during incubation to detect ROP.

A blood sample was taken from each neonate to determine serum IGF-1 and VEGF levels.

Patients and MethodsPatients and MethodsMethodology:

On admission

Every patient was meticulously examined as follows:

– Detailed history taking– Full clinical examination of the studied neonate– Required investigations– Ophthalmic examination: It was performed during

incubation by an ophthalmologist of the NICU to detect ROP and again at 4 to 6 weeks postnatal age for follow up of premature cases.

Patients and MethodsPatients and MethodsMethodology:

After dischargeAll babies were seen at the follow up pediatric clinic at 4 to

6 weeks post natal age to perform the following :

– Routine general and systemic examination – Ocular examination: ROP diagnosis was done by

examination of the fundus with indirect ophthalmoscopy using scleral depression and documentation of the findings with wide angle digital fundus photography Retcam(2). Then recommendation for treatment was assigned to each patient

Patients and MethodsPatients and Methods

– Cytokine analysis: Venous blood samples were collected at 4 to 6 weeks postnatal age (at the time of the ocular examination for ROP) for estimation of IGF-1 and VEGF levels in serum by ELISA techniqe.

– All the cases of the study followed the rules of Medical Ethical Committee of National Research Center.

ResultsResultsSex distribution of cases and controls

Sex distribution of cases (Stage 1 and 2 versus stage 3 and 5)

Stage 1&2 Stage 3&5 n=16 n=18

Sex

Number % Number %

P-value*

Male 10 62.5 6 33.3 Female 6 37.5 12 66.7 0.089

ResultsResults

ResultsResults

Comparison of neonatal anthropometric data between cases and control

Factor Cases Controls P-value*

n=34 n=49

Mean ± SD Range Mean ± SD Range

GA (week) 31.6 ± 3.1 26.0-37.0 34.1 ±1.6 30.0-36.0 <0.001*

Birth weight (kg) 1.5 ± 0.6 0.7-3.3 2.1 ± 0.6 1.0-3.2 <0.001*

Wt at exam. (kg) 2.9 ± 1.5 1.2-7.0 2.9 ± 1.1 1.5-6.5 0.823

Length (cm) 45.1 ± 2.7 41.0-49.0 47.4 ± 3.1 42.0-58.0 0.001*

HC (cm) 33.7 ± 3.1 30.0-42.0 34.1 ± 2.5 31.0-41.0 0.819

ResultsResultsNeonatal anthropometric data of cases (stage 1 and 2 versus stage 3

and 5)

ResultsResultsMedian duration of stay in NICU for cases and

controls

ResultsResults

Different Modes Of Oxygen Therapy

Max FiO Average FiO Max PaO Average PaO

mm

Hg

Cases Controls

P-Value = 0.002

P-Value = 0.021

P-Value = < 0.001

P-Value = 0.002

Max FiO2 Average FiO2 Max PaO2 Average PaO2

ResultsResultsComparison of some clinical data between cases

and controls

ResultsResults

Clinical data of cases (Stage 1 and 2 versus stage 3 and 5)

Factor Stage 1&2 Stage 3&5 P-value*

n=16 n=18

Number % Number %

RDS 12 75.0 18 100.0 0.039*

Neonatal Jaundice 3 18.8 0 0.0 0.094

ResultsResultsPercentage distribution of cases among stage 1, 2, 3,

and 5

29.4

17.6

47.1

5.9

0.0

5.0

10.0

15.0

20.0

25.0

30.0

35.0

40.0

45.0

50.0

Stage 1 Stage 2 Stage 3 Stage 5

Per

cen

tage

ResultsResultsComparison of cytokine serum levels between cases

and controls

45.911.5

330.1

987.5

0

100

200

300

400

500

600

700

800

900

1000

Mea

n

IGF-1 (ng/ml)VEGF (pg/ml)

Cases

Control

P-Value = < 0.001

ResultsResults

Cytokine levels of cases (stage 1 & 2 versus stage 3 & 5)

Factor

Stage 1&2 (n=16) Stage 3&5 (n=18) P-value*

Mean ± SD Range Mean ± SD Range

IGF-1 (ng/ml) 13.6 ± 9.2 1.0 – 28.1 9.7 ± 8.5 0.6 – 24.6 0.324

VEGF (pg/ml) 959.1 ± 257.9 680-1350 1012.8 ± 273.6 635-1500 0.784

ResultsResultsMultivariate logistic regression of the effects of the

variables on ROP

  

B S.E. Sig. OR

95% C.I. for OR

Lower Upper

VEGF 0.019 0.006 0.001 1.02 1.01 1.03

Constant -12.502 3.881 0.001 <0.01B = coefficient, S.E. = standard error of the coefficient, Sig. = significance, OR = odd ratio, C.I. = confidence interval

ResultsResults

Sensitivity and specificity values for IGF-1 and VEGF

Cytokine Cut off 95% CI* Accuracy Sensitivity Specificity

IGF-1 22 ng/ml 18.5 -25.8 88.0% 91.2% 85.7%

VEGF 646 pg/ml 565 - 712 96.4% 97.1% 95.9%

* Confidence interval

ResultsResultsCorrelation between IGF-1 and weight at

examination

0

10

20

30

40

50

60

70

80

90

0 1 2 3 4 5 6 7 8

Weight (kg)

IGF

-1

P-Value = 0.003 r = 0.492

ResultsResults

Correlation between IGF-1 and length

0

10

20

30

40

50

60

70

80

90

38 43 48 53 58

Length (cm)

IGF

-1

P-Value = 0.005 r = 0.477

ResultsResults

Correlation between IGF-1 and head circumference

0

10

20

30

40

50

60

70

80

90

28.0 30.0 32.0 34.0 36.0 38.0 40.0 42.0 44.0

Head Circumference (cm)

IGF

-1

P-Value = 0.015 r = 0.421

ResultsResults

Correlations between IGF-1 and data concerning oxygen administration

Factor Significance (2-tailed) Pearson Correlation

Duration of stay in NICU 0.205 0.223

Ventilation duration 0.044 0.348*

NCPAP duration 0.290 0.187

Max. FiO2 0.651 0.080

Max. PaO2 0.259 -0.199

ResultsResults

Correlations between VEGF and neonatal clinical data

Factor Significance (2-tailed) Pearson Correlation

Birth Weight 0.531 -0.111

Gestational Age 0.154 -0.250

Weight at Examination 0.694 -0.070

Head circumference 0.797 -0.047

Length 0.754 -0.057

ResultsResults

Correlation between VEGF and data concerning oxygen administration

Factor Significance (2-tailed) Pearson Correlation

Duration at NICU 0.767 -0.053

Ventilation duration 0.675 0.075

NCPAP duration 0.719 -0.064

Max. FiO2 0.580 -0.098

Max. PaO2 0.796 -0.046

ResultsResults

Correlation between IGF-1 and VEGF

P-values Correlation

Cases 0.343 -0.168

Controls 0.910 -0.017

ResultsResults

Wide angle digital fundus image of the left eye for one of the cases

ResultsResultsWide angle digital fundus image of the left eye for the same case

three days post avastin injection and laser to the retinal periphery

SummarySummary

Several risk factors were found to be associated with the development and severity of ROP.

Data of this study suggest that LBW, small GA, oxygen therapy, long duration of incubation and critical illness of the neonate were risk factors for ROP.

Regarding severity of ROP, neonates having respiratory distress syndrome or those with small head circumference were found to be at higher risk of developing severe ROP.

ConclusionsConclusions

Low IGF-1 (< 22 ng/ml) and high VEGF (> 646 pg/ml) serum levels can be useful as indicators in ROP screening .

.For each unit (1 pg/ml) increase in VEGF value, the chance of having ROP increases by 3%.

IGF-1 and VEGF were not significant risk factors for severity of ROP.

ConclusionsConclusions

Determination of IGF-1 and VEGF serum levels in the 4th week post partum provides a sufficient and reliable prognostic tool and allows the identification of a group of patients at high risk of developing ROP.

IGF-1 is positively correlated with neonatal weight, head circumference, length and ventilation duration.

Take home MessagesTake home Messages

Take home MessagesTake home Messages Screening of all prematures, less than 37 weeks

gestational age, and /or less than 2500 gm birth weight is importance for detection of ROP and early intervention at stages that yield good outcome.

The timing of this screen should be done at the age of 4 to 6 weeks post natal age. We should not wait until the problem is solved, as this might be too late.

Retinal examination should be repeated approximately every two weeks until treatment is required or retinal maturity is reached.

Take home MessagesTake home Messages

The time of retinal screening of premature infant is important to prevent advanced ROP which may lead to infant blindness.

RecommendationsRecommendations

It is recommended that oxygen intake is to be closely monitored to avoid transient or prolonged phases of hyperoxia as recommended by many international guidelines.

All premature neonates at risk, should be routinely screened at 4 weeks postnatal for serum levels of IGF-1 and VEGF as predictive factors for ROP.

Kits for IGF-1 and VEGF should be readily available for routine determinations in labs and not only for research purposes

Thank YouThank You