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Sources of Funding: P50 HD076210, U1 1U01HD074542-01, Frederick J. and Theresa Dow Wallace Fund of the New York Community Trust, the Mr. Robert S. Dow Foundation, Irena and Howard Laks Foundation_x000D_ _x000D_ This work was supported in part by the Urology Care Foundation Research Scholar Award Program and AUA New York Section Research Scholar Fund

Introduction

Klinefelter syndrome (KS) is characterized by the presence of an additional X chromosome (47,XXY) and leads to sertoli cell only (SCO) histological phenotype. Age dependent progressive hyalinization observed in men with KS and SCO is not seen in 46,XY men with SCO. Hyalinization in KS is believed to play a significant role in age dependent decline in sperm recovery during microscopic testicular sperm extraction. The objective of our study was to evaluate if there are genes differentially expressed between men with KS (47,XXY) and men with 46,XY and histological pattern of SCO. Therefore, providing insight on the role that the additional X chromosome contributes to progressive testicular hyalinization in men with KS. _x000D_ _x000D_

Methods

Total RNA was extracted from tissue harvested during microscopic testicular sperm extraction (mTESE) from patients with NOA and KS, and SCO as well as normal testicular tissue. RNA libraries were sequenced on an Illumina HiSeq 2000 platform. Results were mapped to the genome and transcriptome using TopHat (v2.0.8). Cufflinks was then used to quantify the number of reads. RNA Seq data was expressed as FPKMs and normalized using a TMM, JMP genomic was used to identify differentially expressed (DE) transcripts at FDR = 0.001._x000D_ _x000D_

Results

Testicular tissue from 6 patients with KS 47,XXY SCO, 11 with SCO and 10 with normal spermatogenesis were harvested, processed and sequenced. Using a clustering analysis, RNA expression in SCO 47,XXY was most similar to SCO 46,XY. 10,777 genes were found to be DE between SCO and normal controls largely representing genes expressed in germ cells. However when men with SCO 47,XXY were compared to men with SCO 46,XY we identified that only 546 transcripts (5%) were differentially expressed between these two groups despite both having SCO histology. Further analysis demonstrated downregulation of CELA2A and CELA2B as well as PRSS12 in men with SCO and KS as opposed to men with SCO and normal karyotype. These genes are known to code for serine proteases involved in elastin and collagen metabolism. Elastin and Type 4 collagen are known to contribute the characteristic hyalinization of KS tubules. Therefore, loss of these genes in men with KS may explain age dependent progressive hyalinization of tubules observed commonly in KS. _x000D_ _x000D_

Conclusions

Hierarchical clustering of gene transcripts demonstrates that 46,XY and 47XXY men with SCO share very similar expression profiles. Reduced expression of CELA2A, CELA2B and PRSS12 in men with KS may explain age-dependent progressive hyalinization of seminiferous tubules among these men._x000D_

Funding

P50 HD076210, U1 1U01HD074542-01, Frederick J. and Theresa Dow Wallace Fund of the New York Community Trust, the Mr. Robert S. Dow Foundation, Irena and Howard Laks Foundation_x000D_ _x000D_ This work was supported in part by the Urology Care Foundation Research Scholar Award Program and AUA New York Section Research Scholar Fund