Table 1.

Major phenotypes of FSH gain-of-function genetic models.

Model	Promoter/mutation	Major phenotypes	Implications	References
10 kb hFSHβ targeted expression (pituitary)	HFSHB promoter (Transgenic line)	Both: •Sexually dimorphic expression •Gonadectomy resulted in elevated FSH levels in serum, elevated h/m FSHβ mRNA •Treatment with GnRH increases expression 4 to10- fold, which is suppressed by testosterone/ estradiol •Truncation of sequences upstream of 5′ promoter region retained expression of hFSHβ •Truncation of poly-A sequences downstream of 3′ stop codon in exon 3 resulted in complete loss of expression •Replacement of 3′ poly-A sequences with heterologous sequence failed to rescue expression Males: •Castration: decreased FSH levels in pituitary •Castrated + testosterone treatment: suppressed mRNA content and serum FSH levels (more so than normal littermates) •Intact: increased testicular weights in adults •Intact: higher serum testosterone levels Females: •OVX: increased FSH levels in pituitary •OVX + E2 treatment: FSH suppression in pituitary and serum; mRNA suppression •Intact: normal fertility/ litter size/ number of fertilized embryos	Model for study of hFSHβ regulation	(32–35)
Genetic rescue with 10 kb hFSHβ targeted expression (pituitary);	HFSHB promoter, Fshb null genetic background. (Type I rescue; FR-I) (Combination of a Transgenic and a knockout)	Both: •Targeted expression of FSH in gonadotropes Males: •Fertile; restored testes size and structure/ histology, normal sperm count/motility Females: •Fertile (10/10), normal litters, corpora lutea (CL) in rescued ovaries readily apparent	Model to study effects of pituitary gonadotrope-targeted expression of FSH on Fshb null genetic background	(36, 37)
Ectopic FSH (low)	mMT-1 promoter (mouse Metallothionein-1); Fshb null background (Type II rescue, FR-II) (Combination of a Transgenic and a knockout)	Both: •Ectopic expression of FSH Males: •Fertile; restored testes size and structure/ histology, normal sperm count/motility Females: •Partially fertile (3/10), small litters, small antral follicles and corpora lutea (CL) in rescued 2/3 females died postpartum •Thin uteri, folliculogenesis arrested at pre-antral stage in non-rescued mice •Weak expressors themselves were fertile and had no distinguishable phenotypes from normal littermates	Model to study effects of ectopically expressed FSH	(36)
Ectopic FSH (high)	mMT-1 promoter (Transgenic line)	Both: •Infertile •Elevated serum steroid hormone levels (i.e., testosterone, estradiol, progesterone) Males: •Enlarged seminal vesicles, normal testicular size/development •Increased (epididymal) sperm counts •Castration reduced seminal vesicles to size similar to castrated wild-type littermates Females: •Large hemorrhagic/cystic ovaries •Fluid-filled translucent ovaries •Some follicles halted at pre-antral stage, some developed normally •Large/cystic kidneys, abnormal kidney development •Die 6–13 weeks due to urinary tract obstruction	Model to study possible role of FSH (and steroid hormones) in human reproductive diseases	(24)
FSH genetic rescue	Ovine FSHβ (oFshb) promoter; Fshb null background (Combination of a Transgenic and a knockout)	Double N-glycosylation mutant hFSH compared to WT hFSH •Low levels in serum (both) •Readily detectable (mutant) FSH levels in pituitary as subunit monomer but not as FSH heterodimer Males: •Fertile •Lower testes weights •No rescue with regard to testes phenotypic characteristics (tubule diameter, and sperm counts) Females: •Infertile •No estrus cycles •Hypoplastic ovaries and uteri	Model to study possible therapy for FSH ligand deficiency Model to study role of N-glycans on FSH	(36, 38)
Ectopic HFSHB Tg+	Rat insulin II promoter (RIP II) (Transgenic line) (Combination of a Transgenic and a natural mutant background)	Both: •No significant sexual dimorphism in hFSH levels Males: •Similar testosterone levels to controls (non-transgenic hpg) •No correlation between hFSH and inhibin B levels •Strong positive correlation between hFSH and testis size (at high serum hFSH levels) •Disorganized testes development •Minimal but incomplete spermatogenesis (no fully differentiated spermatozoa) Females: •No apparent estradiol response to hFSH •Strong positive correlation between hFSH and inhibin B levels (similar to WT levels) •Strong positive correlation between serum hFSH and ovaries size •Follicle development to type 7 antral follicles, but no corpora lutea found •hpg (Tg+ and non-Tg) body weights lower than WT controls at 9–11 weeks •hpg Tg+ ovaries 4x weight increase (compared to non-Tg) •Primordial follicle numbers 2 times higher than in WT and hpg non-Tg controls •Secondary follicles restored to normal levels (9–11 weeks) •Number of total antral follicles restored to normal levels (9–11 weeks) •Strong positive correlation between inhibin B and antral follicle count •Increased inhibin A expression (compared to non-Tg) •Dose dependent increase in bone mass (hpg and non-hpg) •Positive correlation between FSH levels and osteoblast/bone surface area •Negative correlation between FSH levels and osteoclast/bone surface area •Ovariectomy of hpg mice resulted in: •[-] decrease of serum levels of inhibin A and testosterone •[-] 47% reduction in bone mass •[-] uterine weights similar to control in non-hpg Tg mice •Uterine weights increased compared to control in hpg (still lower than non-hpg) •No detectable Fshr mRNA in bone cells •Age-specific decline in litter production in females due to increased embryo-fetal resorptions without affecting the number of ovulations.	Model to study FSH actions alone	(30, 39–41)
FSH re-routed	HFSHB Mut; on Fshb null background (Combination of a Transgenic and a knockout)	Both: •Sexually dimorphic expression •Dense core granule and chaperone proteins co-localized in pituitary with mutant hFSHβ (similar to LHβ) •Secretion of mutant hFSH increased 2–4 times in response to GnRH agonist (no significant release in control mice) •Lower levels of serum LH in mice expressing mutant hFSH (comparable Lhb mRNA levels to those in pituitaries of Lhb +/− mice) Males: •No specific phenotypes Females: •Ovarian and uterine morphology similar between mutant and wt hFSHβ expressing mice •Aromatase levels restored to normal in mutant and wt hFSHβ expressing mice at 9 weeks (on Fshb null background) •High progesterone levels in mice expressing mutant FSH •6 times more ovulations in mutant FSH (compared to wt hFSH and normal controls) •Identical primordial follicle counts across all groups •Increased pre-antral follicles, CLs, and follicle size in mutant hFSH-expressing mice •Decreased occurrence of atresia in mutant hFSH- expressing mice •Granulosa pro-survival as well as FSH and LH-responsive genes upregulated in mutant hFSH-expressing mice •Lhb-null mice not rescued by mutant FSH	Model to study differences in secretion patterns of LH and FSH	(37)
FSH Tg+ in milk	Rat β-casein promoter (Transgenic line) Bovine β-casein promoter (Transgenic line)	Males: •No phenotypes reported Females: •Recombinant bovine FSH detected exclusively in milk •Larger lumens in mammary glands of β-casein-hFSH Tg mice than wild type controls •hFSH detected in milk fluids and epithelial cells of Tg mice and not in controls •Amount of hFSH detected in milk proportional to transgene copy number •hFSH increased cAMP levels in hFSH-R transfected cells with competitive binding (biologically active) •Transgenic platelet count 2 times more that of WT controls •26.7% of highest-expressing line displayed both breast and ovarian granulosa cell tumors with hemorrhagic cysts •Mouse FSH and progesterone levels of Tg mice higher in all phases of estrus cycle than non-Tg littermates	Model to study ectopic expression of FSH in mammary glands	(42) (43)
Pig FSH Tg	Chinese Erhualian Boar FSHα/β promoter + gene including long range cis-regulatory elements (Transgenic line)	Males: •Boars: TG compared to WT controls: •Serum FSH levels significantly higher •Semen volume, sperm concentration and motility similar •Germ cells per seminiferous tubule increased •Comparable body weights throughout growth •No significant differences in gut microflora or disease markers Females: •Mice: TG compared to WT controls: •Significant increase in litter number •Significant increase in CL number (at 14–28 weeks) •Increased serum levels of endogenous mouse FSH and estradiol •Decreased serum levels of LH and testosterone- •Decreased LH mRNA content •Boars: TG compared to WT controls •Higher serum FSH levels •Higher pituitary FSHβ content •Smaller litter size •Comparable body weights •No significant difference in serum LH and estradiol	Model to study biological effects of pig FSH	(44) (45) (46)
Inhibin-α KO	Inhibin α-subunit gene deletion (Knockout)	Both: •Infertile •Gonadal stromal tumors •Increased serum FSH levels •Die from cachexia-like symptoms Males: •Testicular enlargement/ hemorrhage •Decrease in number of Leydig cells •Decreased spermatogenesis proportional to tumor size Females: •Ovarian hemorrhage •Decreased folliculogenesis proportional to tumor size	Model to understand and study role of Inhibin/ inhibin-α in development as well as its tumor suppressor activity in gonads	(24, 47)
Inhibin-α / FSH double knockout	Inhibin α-subunit and Fshb gene deletion (Double knockout)	Both: •Delayed body weight loss compared to inhibin single mutants •Less severe cachexia in double mutants compared to mice lacking only inhibin Males: •Compared to inhibin single knockouts, double mutants live longer •Testicular tumors in double mutants are less hemorrhagic Females: •Compared to inhibin single knockouts, double mutants live longer •Ovarian tumors are less aggressive •Folliculogenesis is not disrupted at early stages but eventually hemorrhagic ovarian tumors develop	Model to study how FSH acts as a modifier factor to regulate gonadal tumors in the absence of inhibin	(24)
FSHR gain of function	Rat androgen binding protein promoter (rABP) on hpg background (Combination of transgenic and natural mutation) Asp567Gly mutant when compared to hpg non-Tg littermates: TghFSHRwt TgD567G mutant Constitutively active FSHR mutants; Human AMH promoter driving separately expression of mFshr D580H or D580Y cDNA transgenes; or a D580Y knock-in mouse Fshr allele Constitutively active FSHR on Lhr null background	Males: •Fertile (on non-hpg background) •Infertile (on hpg background) •Testis weights increased nearly 2 times •Treatment with testosterone resulted in larger testis •Testis contained small numbers of both round and elongated spermatids, mature Sertoli cells •Increased number of seminiferous tubules (compared to Tg-FSH group) •Slight rise in serum and significant rise in intra-testicular testosterone levels Compared to non-Tg hpg littermates •overexpression: •No effect on testis weight/serum testosterone levels •No additive effect on testis weight with testosterone treatment •No change in expression of steroid synthesis genes •No changes in testis structure/cellular morphology •Treatment with FSH increased cAMP levels 2 times more, basal levels remained the same •No TSH or hCG binding •2 times increase in testis weight •Synergistic effects on testis weight with testosterone treatment •Increased expression of steroid synthesis genes •Later stage spermatogenesis/ post-meiotic elongated spermatids •Treatment with FSH increased cAMP levels (40% as much as TghFSHwt), basal levels increased two times more •Binds to TSH and hCG (cAMP levels increased by 40% that of FSH stimulation) •Transgenic Fshr D580H female mice demonstrated hemorrhagic and cystic ovaries, loss of immature follicles, increased granulosa cell proliferation, increased E2 production, unruptured and luteinized follicles and occasional teratomas •Most severely affected transgenic Fshr D580H female mice, in addition, displayed increased prolactin levels and mammary gland hyperplasia, pituitary adenoma formation and adrenal defects •Transgenic and knock-in Fshr D580Y mice showed milder ovarian phenotypes with only hemorrhagic cysts Compared to WT males •Fertile •Delayed puberty •Mating trials had lower frequency of pregnancy and litter size •20 times more of Fshr mRNA •40% of serum T levels •Normal spermatogenesis and testis/seminal vesicle size •Treatment with antiandrogen had no effect on spermatogenesis or testis size (though reduced seminal vesicle size) while both were arrested in WT Females: •fertile (on non-hpg background) •No significant differences in ovarian weights between hpg Tg and non-Tg littermates	Model to study downstream pathways involving FSHR signaling	(48, 49) (50) (51)