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The Effect of Erythropoietin on Ischemia/Reperfusion Injury after Testicular Torsion/Detorsion: A Randomized Experimental Study

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Abstract

This study was conducted to investigate the protective effect of erythropoietin (EPO) on ischemia/reperfusion related changes after testicular torsion/detorsion. In a randomized experimental trial 30 male rats were randomly allocated into six equal groups of five rats each. Group I (orchiectomy for histopathologic examination), group II (sham operation), group III (torsion for 2 hours, and ischemia/detorsion for 24 hours, and orchiectomy); group IV (torsion for 2 hours, ischemia/detorsion for 24 hours with erythropoietin injection then orchiectomy), group V (torsion for 2 hours and detorsion and EPO injection and orchiectomy 1 week later, group VI (torsion for 2 hours/detorsion and orchiectomy 1 week later). Two groups (groups 4 and 5) received different protocols of erythropoietin administration after testicular torsion/distortion. other groups were not receiving erythropoietin. Johnsen’s spermatogenesis scoring method and Cosentino’s histologic staging method were used to assess main outcome measures of the study. After the experimentation, Johnsen’s score in EPO Groups was statistically different from the score in some groups not receiving erythropoietin. Cosentino’s score in EPO groups was statistically different from the score in all groups not receiving erythropoietin. Neovascularization, vascular necrosis, vascular congestion, edema, hemorrhage, and acute inflammation were observed in some groups. This study shows short-term protective efficacy of erythropoietin on rat testicular injury after ischemia/reperfusion.

1. Introduction

Testicular torsion is the twisting of the spermatic cord, which cuts off the blood supply to the testicle and surrounding structures. It is more common during infancy and early adolescence. It is a very painful condition, but mainly it is a surgical emergency because it may result in the loss of the affected testicle if not treated promptly. Torsion is the most common cause of testicle loss in adolescent males. Some men may be predisposed to testicular torsion as a result of inadequate connective tissue within the scrotum. However, the condition can result from trauma to the scrotum, particularly if significant swelling occurs. It may also occur after strenuous exercise or may not have an obvious cause [1].

Surgery is usually required and should be performed as soon as possible after symptoms begin. If surgery is performed within 6 hours, most testicles can be saved. Testicular torsion and also the detorsion procedure induce morphological as well as biochemical changes caused mostly by ischemia/reperfusion injury in the testicular tissue [2].

Dysfunction induced by free radicals is the major component of ischemic process due to torsion in the testis, and many studies focused on protective effects of special medications and erythropoietin [3, 4]. Many Animal studies have discovered a protective role for erythropoietin against the aftermaths of ischemia/reperfusion in various organ tissues including kidney, heart, liver, and ovary tissues in animals and nervous system [511]. Few studies have recently been conducted on testicular tissues either [1215]. However, available knowledge is limited considering the methodological variations. The aim of this study was to investigate the protective effect of erythropoietin on ischemia/reperfusion related changes after testicular torsion/detorsion.

2. Materials and Methods

A study was conducted in 2008 in Urology Department of Imam Reza University Hospital in Tabriz, Iran. Animals were 200–300 gr weighted male wistar rats that were breeded in Razi institute of Iran and kept, prior and through the investigation process, under normal and similar light, temperature, and feeding plan in the animal lab in Drug Applied Research Center, Tabriz University of medical sciences. In a randomized experimental trial 30 rats were randomly allocated into six equal groups of five rats each. Both testes of each rat were studied. The groups were treated as follows.

Group 1. Both testes of the five studied rats were orchidectomized and sent for histopathology examination.

Group 2. All five rats underwent lower abdominoscrotal incision (because a wide inguinal canal the testes move freely between abdomen and scrotum) and orchiopexy. After 24 hours both testes of the studied rats were orchidectomized and sent for histopathology examination.

Group 3. All five rats underwent lower abdominoscrotal incision. They received 720 degrees bilateral torsion for two hours (based on other studies) then detorsion and orchiopexy were done on them [12, 16]. After 24 hours both testes of the studied rats were orchidectomized and sent for histopathology examination.

Group 4. All five rats underwent lower abdominoscrotal incision. They received 720 degrees bilateral torsion for two hours then detorsion and orchiopexy were done on them. Then erythropoietin was injected and after 24 hours both testes of the studied rats were orchidectomized and sent for histopathology examination.

Group 5. All five rats underwent lower abdominoscrotal incision. They received 720 degrees bilateral torsion for two hours and then detorsion and orchiopexy were done on them. Then erythropoietin was injected, and after one week both testes of the studied rats were orchidectomized and sent for histopathology examination.

Group 6. All five rats underwent lower abdominoscrotal incision. They received 720 degrees bilateral torsion for two hours, and then detorsion and orchiopexy were done on them. After one week both testes of the studied rats were orchidectomized and sent for histopathology examination.

Prior to incisions general anesthesia was induced using 2.5 mg/kg midazolam as intraperitoneal injection. Erythropoietin was administered intravascular in a dose of 3000 u/kg in groups 4 and 5. The pathologist and evaluator were blind to the type of intervention. Spermatogenesis was assessed using Johnsen’s spermatogenesis scoring method which is based on the concept that testis damage causes a successive disappearance of the most mature germ cell type [17]. Following is the scoring grades:

score 10: complete spermatogenesis with regular tubules;
score 9: many sperms, irregular germinal epithelium;
score 8: few sperms;
score 7: no sperms, many spermatids;
score 6: few spermatids;
score 5: no sperm or spermatids;
score 4: few spermatocytes;
score 3: presence of spermatogonia;
score 2: presence of Sertoli’s cells;
score 1: no cells.

Histological assessments were based on Cosentino’s histologic staging method [18].

Data were gathered and analyzed using SPSS version 15 statistical software package. Measures of continuous scales were compared using Kruskall-Wallis test followed by Mann-Whitney U test as post hoc. Categorical variables were assessed using contingency tables and chi-squared or Fisher’s exact tests. P < 0.05 was considered a statistical significance level in primary tests. Bonferroni correction was used for qualitative parameters in multiple comparisons.

Study was approved by committee of ethics in Tabriz University of medical sciences. Code of ethics on working with lab animals was followed in this research and interdisciplinary principles and guidelines for the use of animals in research were considered.

3. Results and Discussion

Median Johnsen’s score was the highest among rats in group 5 when compared to other rats except control rats in groups 1 and 2 that had not undergone testicular torsion. Mean (SD) Johnsen’s score is compared among the groups in Figure 1. Medians of Johnsen’s score were 10, 10, 2, 3.5, 7.5, and 5 in groups 1–6, respectively.

Figure 1

Mean Johnsen’s score compared among the study groups.

In pairwise statistical assessment of comparisons, it was found that Johnsen’s score in group 5 was statistically different from the score in all groups not receiving erythropoietin. However, Johnsen’s score in group 4 was statistically different from the score in groups 1 and 2 not receiving erythropoietin. Further details are given in Table 1.

Table 1

P values for pairwise statistical comparisons of median Johnsen’s score and Cosentino’s score in groups of rats receiving erythropoietin versus control groups.

Mean (SD) Cosentino’s score is compared among the groups in Figure 2. Medians of Cosentino’s score were 1, 1, 3, 2, 2, and 3 in groups 1–6, respectively.

Figure 2

Mean Cosentino’s score compared among the study groups.

In pairwise statistical assessment of comparisons, it was found that Cosentino’s score in group 5 and 4 was statistically different from the score in all groups not receiving erythropoietin. Further details are given in Table 1.

Statistical assessment of comparisons made between erythropoietin groups and other four control groups found that the frequency distribution in vascular congestion was statistically different only when group 4 was compared with group 1 (P = 0.03); the difference in frequency distribution of neovascularization and vascular necrosis was not statistically significant for any of the comparison pairs; groups 4 and 5 had significantly different frequency of edema only with groups 1 and 2 (P = 0.001); hemorrhage frequency was different when comparing group 4 with groups 1 and 2 (P = 0.001), and also the frequency difference of hemorrhage between groups 5 and 3 was also statistically significant (P = 0.001); groups 4 and 5 had significantly different frequency distribution of acute inflammation only with groups 1 and 2 (P = 0.001) (Figure 3).

Figure 3

Relative frequency distribution of vascular congestion, edema, hemorrhage, and acute inflammation compared among study groups.

The protective effects of erythropoietin on renal and cardiovascular ischemic injuries are shown earlier [5, 11, 19]. Effect of erythropoietin on gonadal ischemia is also shown to be positive relieving ischemic injuries after ovarian torsion in rat and mouse models [6, 8, 9, 20].

In this study we assessed the effect of erythropoietin on ischemic/reperfusion changes after testicular torsion. The results of present study were indicative of efficacy of erythropoietin in relieving the changes after ischemia/reperfusion when compared with similar control groups that had not received erythropoietin. The study also found that seven days after ischemia/reperfusion both Johnsen’s score and histological grading were significantly better in groups receiving erythropoietin.

The first study on effects of erythropoietin on testes injury was published in 2005 by Dobashi et al. They investigated the effects of rat erythropoietin on spermatogenesis by transferring rat Epo gene into cryptorchid testes by means of in vivo electroporation and found that it may reduce the risk of the germ cell loss caused by cryptorchidism [21]. Later in 2007 Yazihan et al. in their study in a five-group rat study found that erythropoietin has antiapoptotic and anti-inflammatory effects following testicular torsion [14]. Two other studies on rat model showed also the positive effect of erythropoietin on ischemic testis injuries [12, 22]. It was found that erythropoietin can decrease cell damage and apoptosis. Köseoğlu et al. concludes that erythropoietin preserves the intact somniferous tubular morphology, lowers the percentage of necrotic seminiferous tubules, and reduces the histological damage [13]. One study used mouse model to assess effect of erythropoietin, but as darbepoetin α, on ischemia caused by testis torsion finding that it affects histological grading. The mentioned study assessments were done four hours after detorsion [15].The major methodological variations in studies conducted on effect of erythropoietin after gonadal ischemia include gonadal type as ovaries versus testes, animal type as rat versus mouse, drug administration form as oral versus parenteral, assessment timing, and drug administration timing and order such that nearly every study had some exclusive methodological aspects. However, regardless of methodological variations, our findings on efficacy of erythropoietin on aftermaths of gonadal ischemia/reperfusion were consistent with available literature.

4. Conclusions

Based on our findings and available knowledge, it can be inferred that erythropoietin has positive effects on gonadal ischemia/reperfusion injury. Specifically we conclude supporting short-term efficacy of erythropoietin on rat testicular injury after ischemia/reperfusion. However, available knowledge, including our findings, has not yet turned complete. Two major areas of necessary future research could be dose-response studies and studying time variance of erythropoietin efficacy in this regard.

Acknowledgment

The authors wish to express their sincere gratitude to Research Vice Chancellor of Tabriz University of Medical Sciences for financially supporting this project.

References

1. Schneck FX, Bellinger ME. Abnormalities of the testis and scrotum and their surgical management. In: Wein AJ, Kavoussi LR, Novick AC, Partin AW, Peters CA, editors. Campbell-Walsh Urology. 9th edition. Vol. 4. Philadelphia, Pa, USA: WB Saunders; 2007.
2. Saba M, Morales CR, De Lamirande E, Gagnon C. Morphological and biochemical changes following acute unilateral testicular torsion in prepubertal rats. Journal of Urology. 1997;157(3):1149–1154. [PubMed]
3. McCord JM. Oxygen-derived free radicals in postischemic tissue injury. The New England Journal of Medicine. 1985;312(3):159–163. [PubMed]
4. Paschos N, Lykissas MG, Beris AE. The role of erythropoietin as an inhibitor of tissue ischemia. International Journal of Biological Sciences. 2008;4(3):161–168. [PMC free article] [PubMed]
5. Parsa CJ, Matsumoto A, Kim J, et al. A novel protective effect of erythropoietin in the infarcted heart. Journal of Clinical Investigation. 2003;112(7):999–1007. [PMC free article] [PubMed]
6. Ergun Y, Koc A, Dolapcioglu K, et al. The protective effect of erythropoietin and dimethylsulfoxide on ischemia-reperfusion injury in rat ovary. European Journal of Obstetrics Gynecology and Reproductive Biology. 2010;152(2):186–190. [PubMed]
7. Carelli S, Marfia G, Di Giulio AM, Ghilardi G, Gorio A. Erythropoietin: recent developments in the treatment of spinal cord injury. Neurology Research International. 2011;2011:8 pages.453179 [PMC free article] [PubMed]
8. Karaca M, Odabasoglu F, Kumtepe Y, Albayrak A, Cadirci E, Keles ON. Protective effects of erythropoietin on ischemia/reperfusion injury of rat ovary. European Journal of Obstetrics Gynecology and Reproductive Biology. 2009;144(2):157–162. [PubMed]
9. Shishavan MK, Sayyah-Melli M, Rad JS, Roshangar L. The ovario-protective effect of erythropoietin against oxidative damage associated with reperfusion following ovarian torsion in rat. American Journal of Animal and Veterinary Sciences. 2009;107(supplement 2):p. S223.
10. Liu X, Shen J, Jin Y, Duan M, Xu J. Recombinant human erythropoietin (rhEPO) preconditioning on nuclear factor-κ B (NF-κB) activation & proinflammatory cytokines induced by myocardial ischaemia-reperfusion. Indian Journal of Medical Research. 2006;124(3):343–354. [PubMed]
11. Johnson DW, Pat B, Vesey DA, Guan Z, Endre Z, Gobe GC. Delayed administration of darbepoetin or erythropoietin protects against ischemic acute renal injury and failure. Kidney International. 2006;69(10):1806–1813. [PubMed]
12. Bakan V, Çıralık H, Tolun FI, Atli Y, Mil A, Öztürk Ş. Protective effect of erythropoietin on torsion/detorsion injury in rat model. Journal of Pediatric Surgery. 2009;44(10):1988–1994. [PubMed]
13. Köseoğlu B, Yilmaz E, Ceylan K, Uzun E, Bayram I, Hizli F. The protective effect of erythropoietin infusion on testicular torsion/ detorsion: an experimental study. International Urology and Nephrology. 2009;41(1):85–91. [PubMed]
14. Yazihan N, Ataoglu H, Koku N, Erdemli E, Sargin AK. Protective role of erythropoietin during testicular torsion of the rats. World Journal of Urology. 2007;25(5):531–536. [PubMed]
15. Akcora B, Altug ME, Kontas T, Atik E. The protective effect of darbepoetin alfa on experimental testicular torsion and detorsion injury. International Journal of Urology. 2007;14(9):846–850. [PubMed]
16. Ozkan MH, Vural IM, Moralioglu S, Uma S, Sarioglu Y. Torsion/detorsion of the testis does not modify responses to nitric oxide in rat isolated penile bulb. Basic and Clinical Pharmacology and Toxicology. 2007;101(2):117–120. [PubMed]
17. Johnsen SG. Testicular biopsy score count–a method for registration of spermatogenesis in human testes: normal values and results in 335 hypogonadal males. Hormones. 1970;1(1):2–25. [PubMed]
18. Cosentino MJ, Nishida M, Rabinowitz R, Cockett ATK. Histopathology of prepubertal rat testes subjected to various durations of spermatic cord torsion. Journal of Andrology. 1986;7(1):23–31. [PubMed]
19. Foller M, Kaestner L, Straface E, Vogel J. Erythrocytes and erythropoietin. International Journal of Cell Biology. 2011;2011:2 pages.972536 [PMC free article] [PubMed]
20. Patel NSA, Sharples EJ, Cuzzocrea S, et al. Pretreatment with EPO reduces the injury and dysfunction caused by ischemia/reperfusion in the mouse kidney in vivo. Kidney International. 2004;66(3):983–989. [PubMed]
21. Dobashi M, Goda K, Maruyama H, Fujisawa M. Erythropoietin gene transfer into rat testes by in vivo electroporation may reduce the risk of germ cell loss caused by cryptorchidism. Asian Journal of Andrology. 2005;7(4):369–373. [PubMed]
22. Ergur BU, Kiray M, Pekcetin C, Bagriyanik HA, Erbil G. Protective effect of erythropoietin pretreatment in testicular ischemia-reperfusion injury in rats. Journal of Pediatric Surgery. 2008;43(4):722–728. [PubMed]

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A 47,X,+t(X;X)(p22.3;p22.3)del(X)(p11.23q11.2),Y Klinefelter Variant with Morbid Obesity

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Abstract

Klinefelter syndrome is the most common type of genetic cause of hypogonadism. This syndrome is characterized by the presence of 1 or more extra X chromosomes. Phenotype manifestations of this syndrome are small testes, fibrosis of the seminiferous tubules, inability to produce sperm, gynecomastia, tall stature, decrease of serum testosterone and increases of luteinizing hormone and follicle stimulating hormone. Most patients with Klinefelter syndrome are tall, with slender body compositions, and reports of obesity are rare. We report the case of a 35-yr-old man with hypogonadism and morbid obesity and diabetes mellitus. He had gynecomastia, small testes and penis, very sparse body hair and his body mass index was 44.85. He did not report experiencing broken voice and was able to have erections. We conducted a chromosome study. His genotype was 47,X,+t(X;X)(p22.3;p22.3)del(X)(p11.23q11.2). In this case, the patient was diagnosed as Klinefelter syndrome. He showed rare phenotypes like morbid obesity and average height and the phenotype may be caused by the karyotype and the excess number of X chromosome. Further studies of the relationship between chromosomes and phenotype are warranted.

Keywords: Klinefelter syndrome, morbid obesity, karyotype

INTRODUCTION

Klinefelter syndrome is the most common type of genetic cause of hypogonadism and occurs in approximately 1.2-1.53 per 1000 male births. This syndrome is characterized by the presence of 1 or more extra X chromosomes, and the most common karyotype is 47, XXY.1 Phenotype manifestations of this syndrome are small testes, fibrosis of the seminiferous tubules, inability to produce sperm, gynecomastia, tall stature, decrease of serum testosterone and increases of luteinizing hormone (LH) and follicle stimulating hormone (FSH). The deficiency of sex hormone influences the development of metabolic syndrome, obesity, and diabetes mellitus.2 Most patients with Klinefelter syndrome are tall, with slender body compositions, and reports of obesity are rare.3,4 Herein, we report a case of Klinefelter syndrome with diabetes mellitus and morbid obesity.

CASE REPORT

A 35-year-old man who visited the Department of Family Medicine due to obesity, was referred to the Department of Endocrinology for the evaluation of hormonal abnormalities. The patient’s height was 174 cm, his body weight was 135.8 kg, his waist circumference was 137 cm, his hip circumference was 146 cm, and his body mass index was 44.85. He was morbidly obese and had gynecomastia, small testes and penis, and very sparse body hair. He did not report experiencing broken voice and was able to have erections.

The level of complete blood cell count were in normal range, white blood cell count 9270/mcL (Neutrophil/Lymphocyte/Monocyte/Eosinophil/Basophil 62.9/20.6/6.1/8.3/0.8%), hemoglobin 12.6 g/dL and platelet 333000/mcL. Routine chemistry showed no specific abnormality other than oral glucose tolerance test, at 2 hours after test glucose was 224 mg/dL. The patient showed increased levels of serum LH (19.88 mIU/mL) and FSH 29.0 (mIU/mL) and decreased levels of serum testosterone (63.0 ng/dL) upon endocrine study. Both testes were reduced in size on testis ultrasounds and there were no sperm present upon sperm analysis. Under the suspicion of primary hypogonadism, we conducted a chromosome study and the patient was diagnosed with Klinefelter syndrome. His genotype was 47,X,+t(X;X)(p22.3;p22.3)del(X)(p11.23q11.2) (Figs. 1 and ​and2).2). He started treatment with testosterone enanthate injections and was followed up in the outpatient clinic.

Fig. 1

Karyotype of this patient.
Fig. 2

Structure of X chromosome that shows translocation and deletion.

DISCUSSION

About 80-90% of patients with Klinefelter syndrome have 47,XXY karyotype, while 10% have mosaicism. Cases of X chromosome structural changes such as the patient described here make up about 1% of Klinefelter patients.5 Morbid obesity and other characteristics in this patient are not common among Klinefelter patients, and were thought to be related to the X chromosome structural changes. In a previous study, a patient with Prader-Willi phenotype and Xq arm duplication showed Prader-Willi characteristics, even though the methylation of 15 (q11-15) was normal.3 That patient also had chromosome duplication, and we therefore conclude that Xq arm translocations and duplications are associated with morbid obesity.

The number of X chromosomes is also related to phenotype. The phenotypes and severity of symptoms in Klinefelter syndrome patients differ according to the number of X chromosomes.6

Klinefelter syndrome cases with metabolic syndrome or diabetes mellitus have been described in some previous studies, but cases of morbid obesity are rare. Studies of XXY or mosaicism patient phenotypes are, in contrast, common. Patients such as the one described in this case and X chromosome studies have been limited to occasional case reports. Further study of the relationships between chromosome and phenotype are warranted.

Footnotes

The authors have no financial conflicts of interest.

References

1. Bojesen A, Gravholt CH. Klinefelter syndrome in clinical practice. Nat Clin Pract Urol. 2007;4:192–204. [PubMed]
2. Saad F, Gooren LJ. The role of testosterone in the etiology and treatment of obesity, the metabolic syndrome, and diabetes mellitus type 2. J Obes. 2011;2011:pii: 471584. [PMC free article] [PubMed]
3. Gabbett MT, Peters GB, Carmichael JM, Darmanian AP, Collins FA. Prader-Willi syndrome phenocopy due to duplication of Xq21.1-q21.31, with array CGH of the critical region. Clin Genet. 2008;73:353–359. [PubMed]
4. Pramyothin P, Pithukpakorn M, Arakaki RF. A 47, XXY patient and Xq21.31 duplication with features of Prader-Willi syndrome: results of array-based comparative genomic hybridization. Endocrine. 2010;37:379–382. [PubMed]
5. Thomas NS, Hassold TJ. Aberrant recombination and the origin of Klinefelter syndrome. Hum Reprod Update. 2003;9:309–317. [PubMed]
6. Tartaglia N, Ayari N, Howell S, D’Epagnier C, Zeitler P. 48,XXYY, 48,XXXY and 49,XXXXY syndromes: not just variants of Klinefelter syndrome. Acta Paediatr. 2011;100:851–860. [PMC free article] [PubMed]

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Axillary silicone lymphadenopathy secondary to augmentation mammaplasty

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Abstract

We report a case involving a 45-year-old woman, who presented with an axillary mass 10 years after bilateral cosmetic augmentation mammaplasty. A lump was detected in the left axilla, and subsequent mammography and magnetic resonance imaging demonstrated intracapsular rupture of the left breast prosthesis. An excisional biopsy of the left axillary lesion and replacement of the ruptured implant was performed. Histological analysis showed that the axillary lump was lymph nodes containing large amounts of silicone. Silicone lymphadenopathy is an obscure complication of procedures involving the use of silicone. It is thought to occur following the transit of silicone droplets from breast implants to lymph nodes by macrophages and should always be considered as a differential diagnosis in patients in whom silicone prostheses are present.

Keywords: Augmentation, breast prostheses, implant, mammaplasty, silicone lymphadenopathy, ruptured breast implant

INTRODUCTION

During the last four decades, silicone has become one of the most extensively utilized materials for the manufacture of breast implants, mainly because it is non-biodegradable and elicits no or little reaction from human tissue. This wide application of implanted silicone prostheses stems from their biological stability, the long-term preservation of their physical properties, combined with minimal tissue reaction and lack of immunogenicity. In spite of that reputation, side effects associated with the utilization of silicone have been well documented in literature. One uncommon side effect of mammary augmentation is silicone lymphadenopathy, defined as the presence of silicone in a lymph node.[1] This case report describes this obscure complication of silicone breast implantation and discusses thoroughly the challenging diagnostic and therapeutic implications of this clinical enigma.

CASE REPORT

A 45-year-old woman presented to our clinic complaining of a lump, located in the left axilla. Despite having been aware of this lesion for two months, she had not sought immediate medical treatment, until she began to notice intermittent pain in her left axilla and a sensation of heaviness. She had undergone bilateral breast augmentation, using subglandular cohesive gel silicone implants of textured shell surface 10 years ago (Mentor™ – 220 cc each).

On physical examination, there was a relatively mobile, hard and non-tender mass, approximately 3cm in diameter that was located in the left axilla.

Mammography demonstrated an irregular contour of the left implant and a highly radiodense axillary lesion, which corresponded to the palpable mass [Figure 1], while a subsequent breast magnetic resonance imaging (MRI) documented the intracapsular rupture (linguini sign) of the left breast prosthesis, but did not show evidence of silicone leakage from the implants [Figures ​[Figures22 and ​and3].3]. Because the patient denied fine needle aspiration cytology (FNAC), excisional biopsy and frozen section analysis of the mass was proposed in order to confirm the benign nature of the lump. Before the excisional biopsy, the patient was reviewed as an outpatient by the plastic surgeon, who had performed the original augmentation procedure. A combined procedure involving excision biopsy of the left axillary lesion and replacement of the ruptured implant was eventually performed.

Figure 1

Mammography showing irregularity of the contour of the left breast implant and a radiodense mass in the left axilla
Figure 2

Axial magnetic resonance mammography revealing gross disorganization and collapse of the left implant with a positive ‘linguine sign’
Figure 3

Sagittal magnetic resonance mammography demonstrating the collapsed intracapsular rupture of the left implant

On gross examination, small amount of pus-like fluid was seen to surround the ruptured implant. Four enlarged lymph nodes were abundant of clear viscous material, which oozed from the cut surface of the specimen. Subsequent histological analysis identified a histiocytic infiltrate with multinucleated giant cells, vacuoles and refractive material consistent with silicone lymphadenopathy [Figures ​[Figures44 and ​and55].

Figure 4

Histological examination showing lymph node with multinucleated giant cells, vacuoles and refractive material consistent with silicone (Haematoxylin and Eosin staining ×200)
Figure 5

Higher magnifi cation photomicrograph revealing liquid silicone droplets appearing as round vacuoles of varying sizes in lymph node parenchyma (Haematoxylin and Eosin staining ×400)

Follow-up is satisfactory to date and 2 months later she remains well, with complete resolution of her initial postoperative discomfort.

DISCUSSION

Silicone has been used in surgery for over 40 years in breast augmentation. It is composed of dimethylsiloxane polymers, which can result in differing properties according to the variation in their chain lengths and cross-links. Despite its initial reputation as a biologically inert material, it has been related with numerous complications including local and systemic granulomatous inflammatory reactions affecting breast tissue, lymph nodes, joint capsules, heart, liver, and kidneys. Silicone lymphadenopathy involving axillary lymph nodes is an uncommon complication of augmentation mammaplasty.[24]

Silicone particles can migrate through tissues by two distinct mechanisms. The first, following rupture or erosion of a silicone-containing surface and secondly, through continued leakage through an intact surface. The risk of rupture or leakage increases with increasing age of the implant, the site of implantation (retroglandular), the presence of local tissue contractures or symptoms and the type of implant. The average age at rupture varies between studies, but is in the region of 10 to 13 years and it is best diagnosed by MRI. Rupture is usually a harmless complication, which only rarely progresses and becomes symptomatic. When leakage does happen, silicone can cause fibrosis and foreign body reaction, especially when combined with certain fatty acids, resulting in pain and contractures. Once silicone particles have breached the confines of their prosthesis, they may be dispersed through any fibrotic reaction to regional lymph nodes by macrophages in the reticuloendothelial system. The granulomatous reactions may present as lymphadenopathy and, when present in the axilla, malignancy of the ipsilateral breast needs to be excluded.[2,5,6]

The presence of silicone droplets in lymph nodes of patients with breast implants suggests that the transit of various elements, either synthetic or biologic, from breast tissue to lymph nodes via lymphatic channels may have a significant passive component. This passive component may be a crucial determinant in the metastatic process. Silicone migration from breast implants to lymph nodes may therefore represent a model that could be useful in understanding the passive component of metastasis in breast cancer.[7]

The clinical importance of silicone lymphadenopathy has several different facets. In patients who have had post-mastectomy reconstructive surgery using silicone gel breast implants, the differential diagnosis of regional lymph node enlargement should include metastatic breast cancer, as well as silicone lymphadenopathy. In most individuals, who have had cosmetic surgery for breast augmentation, one must also recognize the potential for adverse health effects of silicone migration to regional lymph nodes. The association between silicone breast prostheses and systemic diseases is a highly controversial issue. Till now, most epidemiologic studies, found no association between breast implants and a variety of connective tissue diseases, despite the fact that Brown et al. have published a statistically significant link between ruptured silicone gel implants and fibromyalgia, as well as other autoimmune diseases.[8] On the other hand, there are numerous reports of symptoms in women with breast implants, including myalgia, arthralgia, fatigue and sleep disorders, but there is no adequate evidence of such a relation in the literature. Furthermore, the role of silicone in the development of lymphoma deserves mention, since there are several case reports describing primary breast lymphoma in patients with silicone gel breast implants, as well as patients with coexistent silicone lymphadenopathy and lymphoma in the same lymph node.[6,7,9]

In conclusion, silicone lymphadenopathy is a rare complication of procedures involving insertion of silicone-containing prostheses. This case study highlights the fact that patients need a thorough preoperative evaluation with histologic confirmation of the non-malignant nature of regional lymphadenopathy and reinforces the need to employ a high index of clinical suspicion, in order to exclude malignancy, without leading patients to dangerous overtreatment regimes.

Footnotes

Source of Support: Nil

Conflict of Interest: None declared.

REFERENCES

1. Truong LD, Cartwright J, Jr, Goodman MD, Woznicki D. Silicone lymphadenopathy associated with augmentation mammaplasty. Morphologic features of nine cases. Am J Surg Pathol. 1988;12:484–91. [PubMed]
2. Adams ST, Cox J, Rao GS. Axillary silicone lymphadenopathy presenting with a lump and altered sensation in the breast: a case report. J Med Case Reports. 2009;3:6442. [PMC free article] [PubMed]
3. Shaaban H, Jmor S, Alvi R. Leakage and silicone lymphadenopathy with cohesive breast implant. Br J Plast Surg. 2003;56:518–9. [PubMed]
4. Blum A, Abboud W, Shajrawi I, Tatour I. Prolonged fever due to silicone granulomatosis. Isr Med Assoc J. 2007;9:121–2. [PubMed]
5. Gil T, Mettanes I, Aman B, Taran A, Shoshani O, Best LA, et al. Contralateral internal mammary silicone lymphadenopathy imitates breast cancer metastasis. Ann Plast Surg. 2009;63:39–41. [PubMed]
6. Tabatowski K, Elson CE, Johnston WW. Silicone lymphadenopathy in a patient with mammary prosthesis. Fine needle aspiration cytology, histology and analytical electron microscopy. Acta Cytol. 1990;34:10–4. [PubMed]
7. Katzin WE, Centeno JA, Feng LJ, Kiley M, Mullick FG. Pathology of lymph nodes from patients with breast implants: A histologic and spectroscopic evaluation. Am J Surg Pathol. 2005;29:506–11. [PubMed]
8. Brown SL, Pennello G, Berg WA, Soo MS, Middleton MS. Silicone gel breast implant rupture, extracapsular silicone, and health status in a population of women. J Rheumatol. 2001;28:996–1003. [PubMed]
9. Van Diest PJ, Beekman WH, Hage JJ. Pathology of silicone leakage from breast implants. J Clin Pathol. 1998;51:493–7. [PMC free article] [PubMed]

Articles from Indian Journal of Plastic Surgery : Official Publication of the Association of Plastic Surgeons of India are provided here courtesy of Medknow Publications
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Spermatic Cord Knot: A Clinical Finding in Patients with Spermatic Cord Torsion

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Copyright © 2011 A. Al-Terki and T. Al-Qaoud.
This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

Abstract

Pertinent history taking and careful examination often taper the differentials of the acute scrotum; congruently the ability to diagnose acute spermatic cord torsion (SCT) when radiological adjuncts are not available is highly imperative. This observational study serves to present a series of 46 cases of spermatic cord torsion whereby we hypothesize the identification of a clinical knot on scrotal examination as an important clinical aid in making a decision to surgical exploration in patients with acute and subacute SCT, especially in centers where imaging resources are unavailable.

1. Introduction

Reaching the confluence between clinical findings and imaging adjuncts remains a difficult task in diagnosing spermatic cord torsion (SCT) [1]. Awaiting a radiological diagnosis of SCT in a young patient with a high index of suspicion may lead to unnecessary delay especially in patients presenting in the intermediate stage of torsion, hence rapid assessment is mandatory, and salvaging the affected testis is the ultimate goal within the time window available and the present facilities. Previous studies have demonstrated loss of the cremasteric reflex to be 99% sensitive in patients with suspected torsion [2, 3], however, in the young patient in extreme pain and discomfort, eliciting such sign can be cumbersome.

We present a series of cases, whereby upon clinical examination, SCT manifesting as a palpable cord knot distinct from the upper pole of the testes and epididymal head was observed, delineating the site of torsion of the cord: the spermatic cord knot. It is important to be able to demarcate the junction between the epididymal head and the cord where the knot will be felt. Following palpation of the testicle for lie, size, consistency, and to elicit tenderness, using a bimanual approach, the clinical knot is identified by starting at the epididymis and palpating its body up to the head, proceeding upward to palpate the spermatic cord for a semi-hard nodule, denoting the twisting of the cord (Figure 1).

Figure 1

(a) Bimanual examination demonstrating normal findings. (b) Palpation at the junction of the spermatic cord and epididymal head whereby the clinical knot can be felt.

2. Methods and Materials: Case Series

Available data from January 2009 to June 2011 on cases of acute scrotal pain presenting to our emergency department at Al-Amiri Hospital, Kuwait, was reviewed. Data on age (in years), duration of symptoms (in hours), site of pain, ultrasound use, presence of clinical knot on exam, and operative findings were extracted. The primary outcome was the presence of the spermatic cord knot on examination. Descriptive statistics of the series including frequency and percentages is presented stratified by diagnosis. Chi-squared test for trend tests was used to look for an association between age, site of torsion, the operative findings of degrees of rotation of the cord, and the primary outcome. Statistical analysis was conducted using STATA [4].

3. Results

In total, data was available on 114 patients (Table 1): 46 cases of suspected torsion (40%), 32 cases of epididymitits/orchitis (28%), 18 cases of varicocele (16%), 8 cases of inguinal hernia (7%), and 10 cases of undiagnosed pain (9%). The spermatic cord knot sign was seen amongst 40 (87% sensitivity) of the patients with SCT (Table 1), and amongst none of the other patients presenting with other diagnoses.

Table 1

Descriptive statistics of patients stratified by diagnosis.

Amongst patients with SCT, the age range was 4–32 years (mean 18.3 years). The clinical knot sign was observed mostly in patients presenting in the early stage (1 to 7 hours) of SCT (1 to 7 hours: 74%, 7 to 24 hours: 22%, >24 hours: 4%). All patients with suspected SCT were taken for surgical exploration, 44 out of 46 (96%) patients were operated based on the clinical suspicion and finding of the cord knot on examination without the need for supplementary Doppler ultrasound. Most patients were operated within 2 hours of presentation, and contralateral orchiopexy was performed simultaneously; 4 cases (8%) had an unsalvageable testis (Table 2). Ultrasound was performed for two patients whom had presented at a late stage (>24 hours). Most patients had at least a 360-degree rotation of the testicle around its axis (45 patients, 98%, Table 2). Chi-squared test for trend demonstrated a significant association between degree of rotation and presence of clinical knot sign on examination (P = 0.006), however, chi-squared tests did not show an association between age (<16 versus >16 years) and site of torsion (right versus left) with presence of the clinical knot sign on examination (P = 0.81 and P = 0.55).

Table 2

Clinical and operative findings of patients with spermatic cord torsion.

4. Discussion

Our modest series points to the potential aid the clinical knot sign adds to the emergency, pediatric, surgical, and urological staff attending to the case of acute scrotum presenting in the acute and subacute stage, when imaging is unavailable or delays action. Diagnosing SCT can be difficult, and distinction of the scrotal contents is necessary while paying particular attention to identifying the epididymis and delineating the cord from the epididymal head. A common clinical diagnostic dilemma in patients with acute scrotal pain is the inability to differentiate SCT from epididymitis and/or orchitis [5]. We demonstrated that one could help differentiate that by identifying clinical knot sign that was not present in other cases of acute scrotal pain, without delaying surgical exploration.

Earlier, MR imaging has shown specific signs that help differentiate SCT from epididymitis: the whirpool/twisting pattern and the torsion knot, which appear as swirls centered over a low-signal-intensity focus [6]. Later, a report on two cases was published demonstrating similar findings on sonography [7], whereby a central echogenic focus was seen correlating to the low-signal-intensity focus seen on MR equivalent to the torsion knot. Although previous reports have demonstrated the identification of the whirlpool pattern and torsion knot [610], previous literature has not approached this sign on clinical examination. Despite Doppler ultrasound having a high sensitivity and specificity [11] in detecting testicular torsion with blood flow patterns to help delineate torsion from inflammation (epididymitis/orchitis), and alternative techniques such as scintigraphy and MR imaging achieving even higher diagnostic accuracy [12], the use of imaging as an adjunct may only be justified in patients with a low suspicion of acute SCT. Ultrasound was used in our series as an adjunct only for 2 patients with SCT, whom presented in the late stage whereby further delay awaiting imaging would cause no further harm than already present.

Since our aim from this observational study, based on a case series, was to emphasize on a clinical finding, the spermatic cord knot, as a potential adjunct to ultrasound and imaging in centers where these facilities are unavailable, inherently our description lacks comprehensive statistical analysis. In an attempt, our results demonstrate that amongst those without a positive sign on exam, the clinical knot was still evident on surgical exploration, pointing to the difficulty that can be faced in eliciting such sign, and yet a considerably high sensitivity of 86%, and a very low specificity. However, this must be weighed against the small series presented and the fact that all patients taken for surgical exploration had underlying torsion, that is, no true negatives to serve as a numerator for a predictive value of a negative examination. As one would expect, our analysis shows a significant association between degrees of rotation of the testicle around its cord and the presence of knot on examination, however, no association was found between age and site of torsion with presence of the knot on examination.

5. Conclusion

We claim the identification of the clinical knot sign on examination helps to reassure the examining doctor of his/her suspicion of SCT in the acute and subacute stage, most importantly avoiding delay in awaiting imaging findings and decision to surgical exploration. The description of this clinical sign is particularly important to rural centers of limited resources, and in centers where Doppler and MRI studies are not readily available to aid diagnosis. However, as a result of the small number of cases, an inherent limitation of this descriptive series is our inability to reach a firm inference yet, and despite advocating the identification of this sign as a strong suspicion to proceed to scrotal exploration, a larger prospective study would enrich statistical power and serve to calculate more robust estimates of incidence, sensitivity, and specificity, and further facilitating exploration of factors associated with the spermatic cord knot while simultaneously accounting for possible confounders.

References

1. Prando D. Torsion of the spermatic cord: sonographic diagnosis. Ultrasound Quarterly. 2002;18(1):41–57. [PubMed]
2. Kadish HA, Bolte RG. A retrospective review of pediatric patients with epididymitis, testicular torsion, and torsion of testicular appendages. Pediatrics. 1998;102(1):73–76. [PubMed]
3. Rabinowitz R. The importance of the cremasteric reflex in acute scrotal swelling in children. Journal of Urology. 1984;132(1):89–90. [PubMed]
4. StataCorp. Stata/IC 10.0 for Windows. College Station, Tex, USA: StataCorp LP.; 2007.
5. Mushtaq I, Fung M, Glasson MJ. Retrospective review of paediatric patients with acute scrotum. ANZ Journal of Surgery. 2003;73(1-2):55–58. [PubMed]
6. Trambert MA, Mattrey RF, Levine D, Berthoty DP. Subacute scrotal pain: evaluation of torsion versus epididymitis with MR imaging. Radiology. 1990;175(1):53–56. [PubMed]
7. Maroto A, Serres X, Torrent N, Figueras M, Hoyo D, Velayos A. Sonographic appearance of the torsion knot in spermatic cord torsion. American Journal of Roentgenology. 1992;159(5):1029–1030. [PubMed]
8. Kass EJ, Lundak B. The acute scrotum. Pediatric Clinics of North America. 1997;44(5):1251–1266. [PubMed]
9. Mattrey RF, Steinbach GC. Ultrasound contrast agents: state of the art. Investigative Radiology. 1991;26:S5–S11. [PubMed]
10. Mattrey RF. Magnetic resonance imaging of the scrotum. Seminars in Ultrasound CT and MRI. 1991;12(2):95–108. [PubMed]
11. Kravchick S, Cytron S, Leibovici O, et al. Color doppler sonography: its real role in the evaluation of children with highly suspected testicular torsion. European Radiology. 2001;11(6):1000–1005. [PubMed]
12. Wu HC, Sun SS, Kao A, Chuang FJ, Lin CC, Lee CC. Comparison of radionuclide imaging and ultrasonography in the differentiation of acute testicular torsion and inflammatory testicular disease. Clinical Nuclear Medicine. 2002;27(7):490–493. [PubMed]

Articles from Advances in Urology are provided here courtesy of Hindawi Publishing Corporation
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Using Information Technology and Social Networking for Recruitment of Research Participants: Experience From an Exploratory Study of Pediatric Klinefelter Syndrome

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Monitoring Editor: Gunther Eysenbach
Reviewed by Malcolm Koo, Ibrahim Adeleke, and Shirley Williams
Sharron Close, MS, CPNP-PC, PhD,corresponding author1 Arlene Smaldone, CPNP-PC, DNSc,2 Ilene Fennoy, MPH, MD,3 Nancy Reame, MSN, PhD,2 and Margaret Grey, RN, DrPH1

Abstract

Background

Recruiting pediatric samples for research may be challenging due to parental mistrust of the research process, privacy concerns, and family time constraints. Recruitment of children with chronic and genetic conditions may further complicate the enrollment process.

Objective

In this paper, we describe the methodological challenges of recruiting children for research and provide an exemplar of how the use of information technology (IT) strategies with social networking may improve access to difficult-to-reach pediatric research participants.

Methods

We conducted a cross-sectional descriptive study of boys between the ages of 8 and 18 years with Klinefelter syndrome. This study presented unique challenges for recruitment of pediatric participants. These challenges are illustrated by the report of recruitment activities developed for the study. We reviewed the literature to explore the issues of recruiting children for research using conventional and IT approaches. Success rates of conventional recruitment approaches, such as brochures, flyers in medical offices, and physician referrals, are compared with IT-based outreach. The IT approaches included teleconferencing via a Klinefelter syndrome support group, services of a Web-based commercial recruitment-matching company, and the development of a university-affiliated research recruitment website with the use of paid advertising on a social networking website (Facebook).

Results

Over a 3-month period, dissemination of over 150 recruitment brochures and flyers placed in a large urban hospital and hospital-affiliated clinical offices, with 850 letters to physicians and patients were not successful. Within the same period, face-to-face recruitment in the clinical setting yielded 4 (9%) participants. Using Web-based and social networking approaches, 39 (91%) agreed to participate in the study. With these approaches, 5 (12%) were recruited from the national Klinefelter syndrome advocacy group, 8 (19%) from local and teleconference support groups, 10 (23%) from a Web-based research recruitment program, and 16 (37%) from the university-affiliated recruitment website. For the initial 6 months, the university website was viewed approximately 2 to 3 times per day on average. An advertisement placed on a social networking site for 1 week increased website viewing to approximately 63 visits per day. Out of 112 families approached using all of these methods, 43 (38%) agreed to participate. Families who declined cited either travel distance to the study site (15, 22%) or unwillingness to disclose the Klinefelter syndrome diagnosis to their sons (54, 78%) as the reasons for nonparticipation.

Conclusions

Use of Web-based technologies enhances the recruitment of difficult-to-reach populations. Of the many approaches employed in this study, the university-affiliated recruitment website supported by a Facebook advertisement appeared to be the most successful. Research grant budgets should include expenses for website registration and maintenance fees as well as online advertisements on social networking websites. Tracking of recruitment referral sources may be helpful in planning future recruitment campaigns.

Keywords: patient recruitment, research subject recruitment, health information technology, social networking, Klinefelter syndrome

Introduction

Recruiting children for research can present many challenges due to parental mistrust of the research process, privacy concerns, and family time constraints [1,2]. Children with chronic and genetic conditions may further complicate the recruitment process [3]. For studies conducted in the United States, additional challenges exist including regulations and guidelines that direct how researchers contact and enroll participants for studies.

Prior to 1996, medical treatment of children was based on clinical trials and the testing of products and medications that were conducted in adults [4]. Although many treatments were effective for adults, some were shown to be ineffective or harmful to children [4]. In 1996, members of a joint workshop of the American Academy of Pediatrics and the National Institute of Child Health and Development issued a consensus statement calling for children to receive adequately tested treatments recommending efforts to include children in research [4]. In1998, the National Institutes of Health (NIH) established a policy with guidelines requiring that children must be included in all human subjects research that is conducted or supported by the NIH unless there was a scientific or ethical reason for not doing so [5]. The goal of this policy was to increase child participation in research for the purpose of generating data specific to the treatment of children [4,5]. However, inadequate representation of children in selected areas of low-prevalence diseases, orphan conditions (such as Klinefelter syndrome), and genetic conditions persist [6]. More innovative approaches to the enrollment of pediatric volunteers in clinical studies are needed.

The general public increasingly uses information technology (IT) as a source of health information. Approximately 80% of the American public search for health information using Internet sources [7]. Researchers are now turning to the Internet as a tool to recruit target study populations [8-14] for Internet-based interventions in conditions such as hypertension [15], diabetes [16-18], smoking cessation [8,12,19], human immunodeficiency virus risk management [13], and depression [14].

The Internet offers many opportunities for informing potential research participants about a study. These include email [8,20,21], discussion boards, blogs [8], search engines [20,21], study websites [20-22], and Web-based platforms for matching researchers with participants. Each form of Web-based communication provides opportunities and challenges for subject recruitment.

On the one hand, exposure of information to vast numbers of Internet users creates an enormous opportunity for visibility and communication with potential research participants. At the same time, the recruitment process can be sabotaged by problems on the Internet, such as emails sent to spam folders [8-10], discussion board and blog administrators blocking content associated with the researcher [9], and poor choice or lack of adequate keywords on study websites that diminish search engine exposure [23]. A key limitation of the use of Internet-based recruitment activities include the inability to reach socioeconomically or educationally disadvantaged groups as well as culturally diverse populations who may lack access to the Internet or familiarity with its use [10]. In the same vein, potential recruits to a study may not be receptive to unsolicited emails, or may not trust the legitimacy of the sender [8]. Clinical investigators encounter special challenges when attempting to recruit children as research volunteers, especially those who have low-prevalence diseases or genetic conditions [6,24].

The purpose of this paper is to describe methodological challenges associated with the recruitment of children as volunteers in research and to discuss how IT may improve access and enrollment of children in research. A case study of our experience in recruiting boys with Klinefelter syndrome for an exploratory cross-sectional study is used to illustrate specific challenges encountered with a difficult-to-recruit pediatric research population and how IT was used to support the enrollment of participants in the study.

Research Recruitment Challenges

Successful enrollment of clinical research participants is both a science and an art. A number of factors, including patients, health care professionals and researchers, structural and organizational entities, and history, interact to compromise or undermine successful enrollment of patient volunteers into a clinical study [1]. Patients may have limited access to research information and might not fully understand the role of clinical research in the advancement of knowledge for drug and behavioral therapy development [1,21]. They may also worry about or mistrust researchers and their institutions due to lack of understanding about the research process or associated risks and benefits [1,21,25]. Patient characteristics, such as culture, language, and religion [1], may further reduce the chance of successful enrollment. Health care providers may play an important role in gaining access to potential participants, but also may represent barriers to such access [1,2,21].

Health care providers in nonacademic settings may have a limited understanding or interest in clinical trials or may have misgivings about academic institutions [1,26]. Community health care providers may also have concerns about losing control over their patient’s care, or losing the patient to another provider [1,26]. Full-time clinicians are frequently pressed for time in caring for patients and may be concerned about the additional administrative workload and lack of administrative support for research activities [1,2,26]. This concern may lead to financial disincentives for clinical providers to become involved in informing their patients about research enrollment opportunities [26]. Researchers themselves sometimes fail to recognize how they may contribute to recruitment and enrollment problems in their own studies. Lack of training and proficiency in communication for the conduct of research with low-literacy populations may lead to misunderstandings between the researcher and potential participant and, in turn, lower response rates of participation [1].

Other barriers facing researchers include lack of attention to the mistrust of the population to be recruited, failure to demonstrate cultural sensitivity, and lack of training in understanding health care disparities in underserved populations [1]. These barriers are often unrecognized by researchers and get in the way when attempting to gather the desired sample. Structural and organizational factors may also be associated with the desire or ability of people to volunteer for research [1]. Researchers need to consider logistic arrangements to facilitate patient participation, such as creating convenient times and locations for study participation.

Communities may also be sensitive about allowing researchers entry into their environment, especially when they perceive that their participation in the scientific efforts does not result in any return or reward at the community level [1]. This concern makes it very difficult for researchers to re-enter the same community or for the community to be approached by other researchers.

The history of disreputably negative research practices persists in the minds of the public and these perceptions may influence the attitudes of potential research volunteers. The awareness of inhumane treatment by Nazi researchers during World War II and the infamous Tuskegee Syphilis Project conducted by the US Public Health Service from 1932 to 1972 [27] may promote overall fear and mistrust about the research process in the minds of many potential research participants.

Levels of Protection that Challenge Research Recruitment

In the United States, several guidelines offer protection to the public with regard to personal health care and participation in research. Public Law 104-191, also known as the Health Insurance Portability and Accountability Act (HIPAA), was enacted to protect the privacy and personal health information of the public [28]. The HIPAA requirements also guide researchers on how to protect the privacy of research participants. Although all health care providers and researchers are required to obey these laws, many members of the public may be apprehensive of the attendant side effects of disseminating private medical information by researchers. Levels of protection, designed to benefit the public, also may impede progress in the timing and accomplishment of recruitment.

Although pediatric researchers are charged with the responsibility of recruiting children for research, several challenges exist in such efforts. Because parents are legally responsible for their children, it is the parent who must be approached for permission for their child to participate. Parents’ willingness to have their child participate in a study may be influenced by their perception of benefits, risks, and barriers to participation [2]. The child must also assent to the activities of the research project. The child’s willingness to participate in the project may depend upon his/her developmental status and any vulnerability related to illness, chronic condition, or communication disabilities. Children may view research participation as a positive experience, including a wish to help others, reward incentives, and the desire to have a fun experience [2]. These positive motivations may be offset by anticipated unpleasantries, such as blood tests, disagreeable medication regimens, or interruptions in their daily lives [29]. The child-recruit is embedded within a family with complex daily schedules often including parental work, school schedules, and sport practices or other extracurricular activities. All members of the family, including the child’s siblings, influence the busy family schedule. Researchers must anticipate and accommodate time commitments of the family as well as considerations for transportation and commute time. Finally, the parents and the child-recruit must be prepared to agree about certain participation risks and unpleasantries such as completing multiple forms and surveys, or medical examinations, including blood collection.

Methods

Case Illustration: A Study of Boys With Klinefelter Syndrome

The exemplar case illustrates our recent experience with recruiting boys with Klinefelter syndrome for participation in a cross-sectional study. Traditional approaches to recruitment fell short of obtaining the desired sample and expanding the approach with IT resulted in a significant gain in enrollment.

Klinefelter syndrome is a genetic condition caused by the presence of an extra X chromosome (karyotype 47, XXY). This condition occurs in an estimated 1 in 450-500 male births [30,31]. Although it is not rare, it is extremely underdiagnosed. Approximately 64% of affected males are not aware of the diagnosis, and of the 36% who are aware, only 10% are diagnosed in childhood [32]. Klinefelter syndrome in adults is associated with androgen deficiency, gynoid distribution of body fat, gynecomastia, small testes, and azoospermia [33,34]. Individuals diagnosed with Klinefelter syndrome during adulthood report childhood developmental delay; speech, language and learning problems; and psychological issues including depression, shyness, aggression, and social interaction difficulties [35,36]. Klinefelter syndrome poses increased health risks throughout the life span, including increased risk for cardiovascular disease, diabetes, and osteoporosis [37]. Diagnosis of Klinefelter syndrome during childhood may represent an opportunity to address both physical and psychosocial health challenges.

Klinefelter syndrome is a misunderstood condition owing to a paucity of research in children, lack of clear clinical guidelines for treatment during life stages, and unfortunate conclusion errors made by early researchers that suggested men with Klinefelter syndrome were at increased risk for criminal behavior [38-40]. As a result, Klinefelter syndrome families may struggle with inadequate information, lack of support, perceived stigma, and uncertainties about their son’s health [41]. Current research focused on boys with Klinefelter syndrome report fairly small sample sizes, ranging from groups of less than 20 [42-44] to the largest reported cohort of 93 [45]. Misunderstandings about Klinefelter syndrome may contribute to reluctance on the part of many men and families of young sons with Klinefelter syndrome to discuss or disclose information about their diagnosis to others [24].

We conducted an exploratory descriptive study to better understand phenotype, biomarkers, and psychosocial health parameters of boys with Klinefelter syndrome between the ages of 8 and 18 years [46]. The study protocol included a physical examination, blood collection for reproductive and cardiovascular biomarkers, and psychosocial health measurements including quality of life, self-esteem, self-concept, and risk for depression. The Columbia University Institutional Review Board approved the protocol for this study. For this exploratory study, sample size was based on a moderate correlation of at least 0.40 between the clinical characteristics and psychosocial variables as observed in studies of health-related quality of life and polycystic ovary syndrome [47,48]. For a correlation of 0.40 with alpha=.05, a total of 46 subjects were required for a minimum power of 80%. No previous studies with a Klinefelter syndrome population studied the relationship between clinical characteristics and psychosocial health. Recruitment was planned with traditional approaches, including contacting patients in a local pediatric endocrine practice; sending letters to pediatricians, pediatric endocrinologists, geneticists, and genetic counselors; and the use of recruitment flyers and brochures placed strategically throughout the medical center. After sending 850 letters, placing 150 brochures and fliers, and approaching 23 families during clinical visits, only 4 boys were recruited in a 3-month period. It became readily apparent that the traditional approach would fail to achieve the minimum sample size of 46 according to our sample size calculation. Thus, a more innovative approach was devised using IT and social networking.

New recruitment strategies included the development of a study website, in-person information sessions, Web-links, teleconferences, and email access to members of a national and several regional Klinefelter syndrome support organizations, as well as registration with a computer platform clinical recruitment-matching service. Each strategy is briefly described subsequently.

Klinefelter Syndrome Study Recruitment Website

A study information and recruitment website [49] was created using the keywords Klinefelter syndrome, KS, boys with KS, and KS phenotype to increase the likelihood that people searching the Internet for information on Klinefelter syndrome might find the website when conducting searches. The website pages provided information regarding the study and its eligibility requirements, study procedures, and how to contact the researcher for further information or enrollment. The study website home page screenshot is shown in Multimedia Appendix 1.

Patient Advocacy Associations

We contacted a national Klinefelter syndrome advocacy association, Knowledge Support & Action (KS&A) [50], who agreed to place information about our study with the study website link on their website. A screenshot of KS&A home page is provided in Multimedia Appendix 2. Regional Klinefelter syndrome support groups with links to the national organization then invited us to give live presentations about our study at their meetings and also agreed to send emails about the presentation and the study to their members. One of the regional groups, the Klinefelter Syndrome Global Support Group (screenshot is shown in Multimedia Appendix 3), offered a monthly parent teleconference. Over a 3-month period, we were able to explain the purpose of the study and to respond to questions regarding our protocol.

Web-Based Clinical Recruitment-Matching Service

RecruitSource is a search engine and computer platform for matching clinical research participants with researchers [51]. A screenshot of the RecruitSource home page can be seen in Multimedia Appendix 4. Researchers can register details about their study and provide eligibility requirements for matching with potential participants.

Patients who might be interested in research participation register their health information via PrivateAccess [52] as shown in Multimedia Appendix 5. This website is a secure Internet registry that enables them to control who can and cannot see all or selected parts of their personal health information. This IT-based platform prescreens the potential participants who give advance privacy directives about their health information and are asked whether they wish to be contacted by a researcher. The incentive for people using this registry is that they can share their personal health information with properly authenticated doctors, researchers, or family members on a secure Internet platform. All contact information is coded and encrypted for privacy. The potential participant gives specific permission to be contacted by the researcher. Once the patient is registered, the researcher receives information about participants who have expressed an interest in being contacted for possible inclusion in the study. This service is provided at no cost to the researcher if the RecruitSource Web link is accessed via a patient advocacy association. In this case, the study was linked to the KS&A organization, a national advocacy association for Klinefelter syndrome [50].

Social Networking

Social networking is often defined by Web-based platforms, such as Facebook and others. Social networking, however, may also include face-to-face and teleconference transactions with groups, audiences, researcher-participant, and participant-participant networking. Participant-participant networking is the central component to the recruitment strategy known as snowballing [53]. We used all these networking processes in our Klinefelter syndrome study. The interlinking of IT-based and face-to-face networking provided an opportunity for multiple modes of information exposure about the study. Midway into recruitment, we decided to conduct a short trial of a Facebook advertisement (ad) as shown by the screenshot in Multimedia Appendix 6. Because we had not anticipated this strategy a priori, funding for advertising was limited. Nevertheless, we wished to observe how a 1-week social networking ad might impact exposure to the study website.

Results

Of 112 families approached, 43 (38%) agreed to participate. The most frequent reasons for families declining participation was nondisclosure of the diagnosis to their sons (54/112, 78%) and geographic distance from the study site (15/112, 22%). Most parents who had not disclosed the diagnosis to their sons feared that their sons would learn of the diagnosis through participation.

Recruitment approaches for the participants in the Klinefelter syndrome study are summarized in Table 1. Recruitment using IT and social networking yielded a greater number of participants (39/43, 91%) compared to use of traditional approaches (4/43, 9%).

Table 1

Number of participants using traditional and information technology with social networking recruitment approaches to the Klinefelter syndrome study (N=43).

Of the 69 families who declined, over one-fifth (15/69, 22%) came from direct clinical contact; almost twice that number (29/69, 42%) declined during support group presentations, and one-sixth (10/69, 16%) declined during the national KS&A meeting. The most frequent reason for decline was parents not wanting their boys to learn of their diagnosis (n=54, 78%) and travel distance to study site (15/69, 22%).

In an effort to boost activity from general Web users, we placed an ad on Facebook. The ad ran for 1 week in June of 2010, targeting a general audience. Impressions are the raw number of times an ad is shown to different Facebook users. The Facebook ad was shown a total of 2,522,169 times. Social impressions reflect the number of times the ad was shown with social context who visited the study webpage. There were 2835 social impressions for this ad resulting in 509 clicks directly to the study website’s home page. At a total cost of $311 for the week’s ad, this represents the researcher’s cost of $0.61 per visit. Prior to placing the ad on Facebook, the study website received 2 to 3 visits per day. During the week of Facebook advertising, website visits climbed to an average of 63 visits per day. The Klinefelter study website activity increase in response to the Facebook ad can be seen in Figure 1.

Figure 1

Response (visits per day to the Klinefelter syndrome study website) during Facebook advertisement period showing increase in activity during Facebook advertising.

Because multiple techniques were employed to attract this difficult-to-reach population, it is difficult to attribute any one recruitment approach to increasing the number of participants in this study. Figure 2 shows a timeline of the 1-year recruitment process.

Figure 2

Recruitment to the Klinefelter syndrome study by source timeline.

Discussion

The Internet represents an increasingly valuable resource for researchers, especially for those who wish to understand the social and cultural context of the populations they are attempting to reach [54]. Since the advent of IT as a common mode of communication, researchers have learned many lessons about the pearls and pitfalls of using this recruitment approach. In difficult-to-reach and vulnerable populations, such as Klinefelter syndrome families, our experience has led us to understand better that more than 1 recruitment technique may be required to inform potential participants and to foster trust in them. We believe that the construction and launch of the study website served these 2 important purposes. We were, however, unable to attribute increased participation due to any 1 technique, including placement of the Facebook ad. Important lessons were learned during this challenging recruitment process, such as the need to track how participants make decisions about whether to participate. We were unable to track which potential research candidates came from the study website activity while the Facebook ad ran because we did not have access to the server log. Anecdotally, several families reported that they chose to participate only after being exposed to the study information from multiple sources. Some families reported that friends or other family members who saw the Facebook ad contacted them to let them know about the study. Once informed, these families conducted either a general Internet search, visited the website directly, or visited the KS&A website for more information. Most importantly, we discovered that we need to track sources of recruitment more carefully in the future by surveying participants about how they found out about the study and also by looking at server logs whenever possible. It would also be helpful to track website visits by Internet protocol (IP) addresses to examine how many potential candidates are first-time or repeat visitors. Because we were unable to attribute which of our recruitment responses came from the Facebook ad, we are unable to estimate the cost per participant. This information would have been very helpful in planning cost allocation for a future study.

Since the advent of IT and social networking in the scientific community, there has been a steady evolution of its use for recruitment and Internet-based interventions. Even within the past 5 to 7 years, much has been learned about the limitations of Internet-based approaches and how such problems might be mitigated.

Although early experience with the use of IT-based recruitment for clinical research, as reported by Koo and Skinner [8], was disappointing, others have offered solutions to optimize challenges that make this form of recruitment difficult. Murray et al [20] solved issues related to mass emailing and spam management by providing recipients with the option to unsubscribe in order to decline further contact by researchers. They were also able to demonstrate the benefits of advertising their study on the home page of a well-known and trusted charity. Our Klinefelter syndrome study recruitment was greatly enhanced by our exposure with the KS&A national advocacy association and with support groups. Ip et al [9] addressed IT recruitment challenges by developing a guide describing a 12-step process to improve visibility and popularity of recruitment messages. The goal of this guide was to increase the interest of potential participants and to offer researchers ways in which to anticipate and respond when IT communication difficulties arise. Recent work with Ramo and Prochaska [11] demonstrated the value of Facebook advertising as an effective mechanism to reach young adults in clinical research. However, reaching a target group under the age of 18 years imposes special issues. For example, although children can be attracted to recruitment advertising for research, they would still be required to obtain parental consent for participation. Although social networking may interest a child about a research project, additional means of informing and developing trust with a parent are still necessary. Sullivan et al [13] and Graham et al [12] each illustrated how changing the composition of banner advertising may improve communication to desired target groups. In the case of pediatric research, such customization may promote discourse between parents and children. The recruitment process, as described by Patel et al [21], is explained as a dialog or discourse that takes place between the investigator and the potential research participant. In the case of minor children, discourse needs to be promoted between investigator, parent, and child if pediatric recruitment is to be successful.

Our recent experience in recruiting boys for the Klinefelter syndrome study can be described as a multilayered strategy of communication using IT. The process of communication began with a traditional print exposure that proved to be ineffective. Adding the various IT communication approaches, including the study website, a computer-based research recruitment website, social networking on Facebook, exposure via support groups online, and by teleconference, offered parents multiple exposures to study information. Although our original sample size calculation called for 46 participants based upon an effect size of 0.40, the effect size from our study proved to be larger (–0.47). We believe that the overall number of participants (43 boys) did not negatively affect the study.

Conclusions

Recruiting boys for a study on Klinefelter syndrome proved to be a challenging endeavor that was best accomplished using IT-based techniques. Important lessons were learned as we dealt with early recruitment challenges. The first lesson is that multiple exposures to the study information and personal contact with the researcher may be helpful in fostering parental trust. Parents must believe that the study, the institution, and the researcher are trustworthy before they will agree to have their child take part. These acts of communication, presented in multiple ways, were central to the success of our recruitment effort and are distinct advantages offered by IT-based strategies. Expenses related to website creation, registration of a domain name, website maintenance, and planning for social networking advertising were not initially anticipated by us, but should be considered by future researchers in the planning process when study budgets are developed. A limitation of our reported recruitment observations is that an in-depth recruitment analysis was not conducted to determine how multiple recruitment exposures occurred. Future IT-based recruitment efforts should preplan the collection of profile data, including IP addresses and tracking of how, when, and how many times a recruitment website was visited. This type of data may assist in the planning of customized approaches for the creation of more effective social networking banner advertising. Nevertheless, the observations from this study may advance the understanding of how difficult-to-recruit participants, like children, might be reached and have parental communication needs met with a view to obtaining their consent to participate in a study. It is noteworthy to mention that there has been inadequate representation of children in Klinefelter syndrome research and in other genetic conditions.

Researchers need to expand their knowledge of how potential recruits might be encouraged to participate in studies by understanding the utility of traditional approaches versus IT and other social networking approaches for recruitment. By offering multiple opportunities for exposure, parents have the opportunity to digest and think about the idea of having their child participate in a study. Because IT and social networking have become well-accepted modes of communication, these tools enable the researcher to layer the recruitment message in order to optimize the likelihood that recruitment efforts will be successful.

Acknowledgments

This research was supported by The Pediatric Endocrinology Nursing Society, the National Association of Pediatric Nurse Practitioners, the Alpha Zeta Chapter of Sigma Theta Tau, Columbia University’s CTSA grant No. UL1 RR024156 from NCATS-NCRR/NIH and Yale School of Nursing T32 Post-Doctoral Fellowship Training Grant, No. 5T32NR008346-08. The content is solely the responsibility of the authors and does not necessarily represent the official views of the NIH.

Abbreviations

IP
Internet protocol
IT
information technology
KS
Klinefelter syndrome

Multimedia Appendix 1

Screenshot of Columbia University Klinefelter syndrome study website.

Multimedia Appendix 2

Screenshot of Knowledge Support & Action (KS&A) Klinefelter syndrome advocacy association website.

Multimedia Appendix 3

Screenshot of Klinefelter Syndrome Global Support Group website.

Multimedia Appendix 4

Screenshot of RecruitSource website.

Multimedia Appendix 5

Screenshot of PrivateAccess website.

Multimedia Appendix 6

Screenshot of Klinefelter syndrome study Facebook advertisement.

Footnotes

Conflicts of Interest:

Conflicts of Interest: None declared.

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