DNA fragmentation severely increased. Furthermore, male factor showed anincrease in T-DNAF and recurrent pregnancy loss presented morphologicallynormal sperm with pathological increased DNA fragmentation. We canconclude that sperm DNA evaluation provides a valuable contribution tothe diagnosis of infertility in the above populations.

Supported by: Institutional.

P-1003 Thursday, October 17, 2013

ELECTROEJACULATION FOR THE TREATMENT OF THE ANE-JACULATORY MALE. S. W. J. Seager. National Rehabilitation Hospi-tal, Washington, DC.

OBJECTIVE:We describe our experience over the last 27 years with Elec-troejaculation (EE) in neurologically impaired men.

MATERIALSANDMETHODS:We use a specially designed electricallyisolated rectal probe with a built-in temperature monitor with a shut-off at apreset temperature. Electrostimulation is supplied for a maximum of tenminutes. Stimulation parameters range from 4-17 volts and 100-500 milli-amps.

RESULTS: From 1990 to date, we have performed EE on 672 anejectorymales and obtained ejaculates in all 672 for a success rate of 100%. 92% ofthe subjects had sperm in their ejaculate. The subjects were men who had spi-nal cord injury (SCI), complications of pelvic surgery, surgical treatment fortesticular cancer, viral infections causing neurological damage, MultipleSclerosis (MS), Spina Bifida, Diabetics, Idiopathic anejaculation caused byneurogenic or psychogenic conditions, failure to produce an ejaculate theday of collection for egg retrieval for IVF or ICSI. A new recent innovationin the use of this method of treatment is in adolescent or teenage boys withnon-Hodgkins disease prior to their treatment of radiation and chemotherapywhich renders them sterile. The mean age range of men with SCI was 32.1years of age (16-61 years). For the non-SCI group it was 38.5 years (25-53years). Sperm was present in 92% or 618 of the men who were ejaculated.No adverse effects were reported.

CONCLUSION: In conclusion, EE is an effective, safe and repeatablemethod of sperm retrieval in neurologically impaired or other males whocannot ejaculate.

P-1004 Thursday, October 17, 2013

ABSTRACT WITHDRAWN

P-1005 Thursday, October 17, 2013

VASCULAR ENDOTHELIAL GROWTH FACTOR (VEGF) POLY-MORPHISM GENOTYPE POSSIBILITIES AND THE ETIOPATHO-GENIC FACTORS OF DNADAMAGE IN SPERM. C. G. Petersen,a,b,c

L. D. Vagnini,b A. L. Mauri,a,b J. B. A. Oliveira,a,b,c R. L. R. Baruffi,a,b

J. G. Franco, Jr.a,b,c aCenter for Human Reproduction Prof. Franco Jr, Ri-beirao Preto, Sao Paulo, Brazil; bPaulista Center for Diagnosis Researchand Training, Ribeirao Preto, Sao Paulo, Brazil; cDepartment of Gynecologyand Obstetrics, Botucatu Medical School -Sao Paulo State University - UN-ESP, Botucatu, Sao Paulo, Brazil.

OBJECTIVE: To analyze whether there are correlations between theVEGF -1154 guanine (G)/adenine (A) polymorphism and etiopathogenicfactors of DNA damage in sperm.

DESIGN: Cross-sectional.MATERIALS AND METHODS: A total of 177 men were included. The

DNAwas extracted from peripheral blood, and the VEGF gene single nucle-otide polymorphisms (SNPs) -1154G/Awere genotyped using real-time PCRwith Taqman Universal PCR Master Mix and Taqman SNP genotyping as-says. At least 200 spermatozoa were examined in each semen sample (oneper subject) and the following spermatozoa percentages were determined:DNA fragmentation (with TUNEL assay), abnormal chromatin packaging/underprotamination (using chromomycin A3/CMA3), apoptosis (using an-nexin-V) and mitochondrial damage (using MitoTracker Green and JC1).All data were evaluated using c2 tests.

RESULTS: VEGF genotypes with A alleles were associated with high per-centages of apoptotic spermatozoa. There was no observed association be-

S438 ASRM Abstracts

tween VEGF genotypes or alleles and DNA fragmentation, protaminationor mitochondrial damage in sperm (Table 1).

TABLE 1. Results

%Mitochondrial

VEGF

%DNA

fragmentation

Vo

%CMA3

positivity

l. 100, No

%Apoptosis

. 3, Supplem

damage

MitoTracker

Green

ent, Septe

JC1

Genotypes

G/G

13.4�6.5 56.0�16.8 18.3�5.4 30.1�18.4 34.4�22.2

G/A

12.6�8.0 54.2�17.3 20.4�6.3 30.4�18.5 34.6�16.6

A/A

10.7�5.0 51.3�15.7 21.8�8.8 20.2�16.3 26.9�14.6

P

0.12 0.50 0.09 0.12 0.35

Genotypes

G/G

13.4�6.5 56�16.8 18.3�5.4 30.1�18.4 34.4�22.2

G/A+A/A

12.1�7.3 53.3�16.8 20.8�7.1 26.6�18.2 31.6�16.1

P

0.06 0.36 0.04 0.33 0.90

Alleles

G

13.1�7.2 55.2�17 19.3�5.9 30.2�18.3 34.5�20

A

12.1�7.3 53.3�16.8 20.8�7.1 26.6�18.2 31.6�16.1

P

0.18 0.47 0.14 0.27 0.65

CONCLUSION: The results indicate that VEGF gene -1154G/A polymor-phisms are related to sperm apoptosis (worse results associated with the Aallele). Further investigations should be conducted to confirm these findingsand to determine their clinical implications.

P-1006 Thursday, October 17, 2013

SPERM CHROMATIN DISPERSION (SCD) TEST IDENTIFIESMORE SPERMATOZOAWITH DNA DAMAGE THAN THE TUNELASSAY INMENWITH UNEXPLAINED INFERTILITY. C. M. Feij�o,S. C. Esteves. Andrology Laboratory& SpermBank, ANDROFERT, Androl-ogy & Human Reproduction Clinic, Campinas, SP, Brazil.

OBJECTIVE: This study aimed at comparing the TUNEL assay and thesperm chromatin dispersion (SCD) test for the determination of spermDNA damage in men with unexplained infertility.DESIGN: Prospective comparative experimental study.MATERIALS AND METHODS: In 15 subfertile men with non-identifi-

able infertility factors according to our routine investigation, sperm DNAdamage was determined manually in the same semen samples using the ter-minal uridine nick-end labeling (TUNEL) assay with fluorescence micro-scopy and the SCD test with bright-field microscopy. The TUNEL assaywas performed using the Apoptosis Detection TACS kit (R&D Systems,USA) while SCD was carried out using the Halosperm kit (HalotechDNA, Spain). A total of 400 cells were evaluated on each slide. Datawere analyzed with Mann-Whitney and Spearman’s rank correlation.ROC analysis was performed using TUNEL as the ‘‘gold standard’’ toselect possibly optimal cutpoints for assessing DNA fragmentation usingSCD.RESULTS: SCD test detected a significantly higher proportion of sperm

with fragmented DNA (20.6% � 14.0%) than the TUNEL assay (11.5% �7.3%; P¼0.001). Receiver operating characteristic analysis revealed that15% was the best SCD cutpoint to classify patients in the same levels ofDNA fragmentation, normal or abnormal, as determined by TUNEL, withan accuracy of 70%. However, Spearman’s rank correlation showed thatthe methods were not comparable (r¼0.29).CONCLUSION: Our data indicate that the SCD test is more sensitive than

the TUNEL assay for the assessment of DNA damage in men with unex-plained infertility. While the TUNEL assay depends on a terminal transferaseto directly incorporate fluorescent UTP at single and double 3’-OH- freeends, SCD involves the combination of DNA denaturation and depletion ofproteins protecting the DNA. Thus, it is important to distinguish betweenthe methods as they deferentially evaluate sperm DNA damage.Supported by: Institutional.

mber 2013