The signi

ﬁ

cance of insulin-like factor 3 as

a marker of intratesticular testosterone

Male factor infertility is a complex clinical problem caused by

multiple contributory factors. Clinicians currently rely on

semen analyses as a tool for diagnosis, an indicator of

spermatogenesis, a barometer of treatment success, and an

indirect predictor of normal intratesticular testosterone

(ITT). The inherent variabilities and

ﬂ

uctuations in semen

analyses, as well as in ITT, has driven researchers to seek

novel biomarkers that may act as more accurate surrogates

of testicular function during treatment for testosterone-

related infertility.

The randomized trial by Roth et al.

(1)

seeks to identify

a serum marker of ITT, which is typically obtained through

an invasive

ﬁ

ne-needle aspiration biopsy. Although the in-

vestigators conclude that insulin-like factor 3 (INSL3) can

be useful in monitoring the ef

ﬁ

cacy of human chorionic

gonadotropin (hCG) stimulation in the hypogonadotropic

infertile male

(1)

, the possibilities for further, more diverse

applications exist. Indeed, given that a normal ITT is

necessary for functional spermatogenesis

(2)

to follow ITT levels with a serum marker would allow cli-

nicians to track testicular function in a minimally invasive

manner throughout the course of male infertility treat-

ments. Furthermore, in patients with hypogonadism and

testosterone-stimulating supplementation, the values of

ITT and testicular function could be monitored in a mini-

mally invasive manner and adjusted to preserve fertility

when desired. These applications are currently limited by

the unknown normal range of ITT and the complicated

physiologic mechanisms that control spermatogenesis;

however, future work will no doubt begin to clarify many

of these questions.

As published in this issue of

Fertility and Sterility,

the

investigators present the results of a prospective, randomized

clinical trial on 37 healthy men (ages 18 to 50 years)

(1)

More speci

ﬁ

cally, this current report is a subanalysis of a pre-

viously reported clinical trial

(3)

. Luteinizing hormone (LH)

and follicle-stimulating hormone (FSH) were antagonized

centrally with acyline (a GnRH antagonist) followed by

randomization to supplementation with either testosterone

gel or various low concentrations of hCG

(1)

. Testicular

ﬁ

ne-needle aspiration for ITT and serum values of testosterone

were obtained before acyline administration and again 10 days

after

complete

gonadotropin

suppression

(1,

The

investigators then examined the effects of their experimental

protocol on several potential serum biomarkers of ITT

including INSL3, 17

-hydroxyprogesterone (17-OHP), anti-

€

ullerian hormone, and inhibin B, with the intent of determin-

ing correlation and statistical signi

ﬁ

cance

(1)

A testicular protein produced by the Leydig cells,

INSL3 is the primary focus of this study and the only

serum marker that the investigators found to increase lin-

early with low-dose hCG therapy

(1)

.Insu

in-

ikefac

tor3

increases during puberty, decreases with age, and is pri-

marily regulated by LH

(1, 2)

. In the clinical context,

they theorized that INSL3 re

ﬂ

ected ITT levels, thus

allowing clinicians to monitor the status of testicular

function during androgen suppression or by serving as

potential

biomarker

spermatogenesis

(2)

Unfortunately, it is dif

ﬁ

cult to conclude that INSL3 can

serve as a biomarker for spermatogenesis because,

although INSL3 is involved in cryptorchidism, the only

known role for INSL3 in adults is in bone metabolism.

Therefore, a Leydig cell origin for INSL3 is accepted and

a relationship to ITT proven

(1)

, but the mechanisms and

reasons for the interaction between INSL3 and the LH/

FSHax

isispu

ive

Indeed, in men with normal hormone serum levels, INSL3

is unaffected by hCG-evoked gonadotropic stimulation

(4)

.It

is only in a chemically induced state of gonadotropin depriva-

tion that INSL3 signi

ﬁ

cantly decreases, and then recovers, in

response to hCG

(4)

: mimicking the changes seen with ITT

(1)

In this experimental perturbation, serum levels of INSL3 were

statistically signi

ﬁ

cantly correlated with serum testosterone

and ITT while the serum levels of inhibin B, antim

€

ullerian hor-

mone, and 17-OHP exhibited no statistically signi

ﬁ

cant cor-

relations

(1)

With respect to the relationship between ITT and INSL3

being statistically signi

ﬁ

cant, it is known that ITT concentra-

tions are quite variable among fertile men and that they typ-

ically re

ﬂ

ect LH pulsatility

(2, 3)

. Indeed, ITT values in

infertile men are completely unknown, and the participants

in their study only exhibited a parallel response when

placed in an arti

ﬁ

cial state of hypogonadotropic gonadism.

This suggests that although INSL3 correlated with ITT in

certain situations, employing INSL3 as a spermatogenic

biomarker in the general population would be premature.

Indeed, neither the normal levels nor the absolute value of

ITT controlling spermatogenesis is known

(2)

. Further

complicating

the

relationship

between

ITT

and

spermatogenesis is that knockout mice models of the LH

receptor still exhibit spermatogenesis, and very low levels

of ITT (

20.2 ng/g) can support spermatogenesis in

humans with a mutated LH gene

(2)

. Moreover, 1% to 2%

of ITT is converted to estradiol by aromatase, which, along

with the interaction of estradiol and FSH

(2)

, yields a much

more complicated description of ITT physiology.

The article by Roth et al.

(1)

presents a subset analysis of

a well-conducted, randomized, placebo-controlled clinical

trial. It discusses a physiologic proof of concept: in patients

with an inhibited gonadotropic axis, INSL3 can serve as

a marker of ITT. Although the

ﬁ

ndings of this study seem

to speci

ﬁ

cally highlight patients with Kallmann

’

s syndrome,

it would be interesting to compare the ITT in patients with

normal spermatogenesis with those who have testicular fail-

ure. Nevertheless, it is tempting to speculate that the rela-

tionship between ITT and, by extension, spermatogenesis

can be quanti

ﬁ

ed by a serum biomarker. Whether or not

INSL3 is such a marker obviously requires further investiga-

tion, and the work by Roth et al.

(1)

paves the way toward

this discussion.

Jason R. Kovac, M.D., Ph.D.

Larry I. Lipshultz, M.D.

Scott Department of Urology, Baylor College of Medicine,

Houston, Texas

VOL. 99 NO. 1 / JANUARY 2013