Testicular Cadmium (Cd) more informative than histology for varicocele prognosis

Abstract

Objective: Our prior analyses of testis biopsies (TB) from infertile men with varicocele showed that [1] heat shock protein 70–1 (HSP 70–1) expression is protective, associated with decreased apoptosis and increased pre-op sperm densities, and [2] that testicular Cd levels > 0.45 ng/mg dry wt are destructive, associated with reduced spermatogenesis and poor response to varicocele surgery. Animal studies show that environmental exposures (e.g., Cd) damage testicular basal lamina (BL). Increased BL thickness occurs early in adolescent varicocele and is more severe in adults. Does HSP 70–1 expression, Cd levels and BL sclerosis correlate in infertile men with varicocele and does degree of sclerosis predict outcome of varicocele repair? Design: Assessment of BL width, Cd levels, HSP 70–1 mRNA expression and apoptosis in bilateral TB of men with left (LV) and with bilateral varicocele (BLV). Controls were TB from obstructive azoospermics (OA). Materials and Methods: [1] IRB-approved bilateral TB by percutaneous needle aspiration (LV, n=7; BLV, n=20; OA, n=5); [2} BL measurements on H&E; stained TB sections with a vernier ocular; [3] Cd levels by atomic absorption; [4] RNA from each TB used in RT-PCR with HSP 70–1-specific primers; and [5] Apoptosis in TB sections by TUNEL. Results: TB were divided into 5 groups: [A] normal (BL ≤ 7 μm), [B] slightly thickened (BL 7.1–10 μm), [C] initial sclerosis (BL 10.1–20 μm), [D] advanced sclerosis (BL 20.1–35 μm), and [E] complete sclerosis (BL >35 μm). OA TB presented with A and some B BL while all varicocele TB contained B-D BL. BL width correlated in matched left and right TB (Pearson correlation [PC], r = 0.5, P <0.04). Degree of sclerosis positively correlated with % apoptosis (PC, r = 0.7, P <0.0001) and negatively with HSP 70–1 (analysis of variance [ANOVA], P <0.04). Though Cd and % apoptosis correlated mildly (PC, r = 0.45, P <0.004), Cd and BL width were unrelated (PC, r = 0.1, P=0.6, NS). BL width was unrelated to pre-op sperm count (PC, r = -0.08, P=0.7, NS) and post-op sperm increase (PC, r = 0.2, P=0.4, NS). LV and BLV did not differ for any parameters. Conclusion: Animal studies indicate heat and/or osmotic stress (both implicated in varicocele pathophysiology) induce HSP 70–1 and HSP 70–1 absence increases apoptosis after osmotic stress. Given that intra-male variability in scrotal temperatures makes it unlikely that heat is the HSP 70–1 inducer with varicocele, these data suggest that increased venous/interstitial pressures cause sclerosis. Sclerosis is not an indicator of spermatogenic potential. Prior studies show Cd alters calcium homeostasis in testis, changing gene expression and mRNA splicing. These changes plausibly explain why elevated TB Cd is a marker for failure to increase spermatogenesis post-op. Support by NIH Grant No. ES10496 to SHB.