LEPTIN DETERMINANTS IN REPRODUCTIVE MEDICINE

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LEPTIN DETERMINANTS IN REPRODUCTIVE MEDICINE

• Moshood Olatinwo, M.D.

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Objectives

• Leptin has significant role in reproduction.
• Leptin is essential for the onset of puberty.
• Leptin directly regulates ovarian responsiveness
  to gonadotropins.
• Leptin directly regulates endometrial receptivity
  to implantation.

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The Effects ofFood Availability on Reproduction?

• Of the many environmental factors that
• can influence a mammals reproduction,
• food availability must be accorded the
• most important role.

F. H. Bronson, Mammalian reproductive
biology. Chicago University of Chicago Press,
1989, p. 88.
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Ingalls et al., J. Hered. 41317-8 (1950)
These animals are obese and infertile
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Parabiosis Experiments
ob/ob normal weight gain of ob/ob mouse
suppressed. db/db normal normal mouse slowly
loses weight and dies of apparent
starvation. db/db ob/ob ob/ob mouse rapidly
loses weight and dies of apparent starvation.
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In 1994, the leptin gene was cloned from the ob
mouse the leptin receptor was subsequently
cloned from the db mouse.
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Appetite/Feeding Metabolism
LH/FSH Reproduction?
Metabolic Signal
Exercise
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What is Puberty?

• Puberty is the stage of physical maturation in
  which an individual becomes physiologically
  capable of sexual reproduction
• Characterized by the activation of the
  hypothalamic-pituitary-gonadal axis
• Profound biological, morphological, and
  psychological changes to which the adolescent is
  subjected

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Response of the Pituitary to Exogenous GnRH
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Changes in Gonadotropins levels at Puberty

• FSH and LH secretion is high in newborn, but
  falls to low levels in few weeks.
• At puberty
• Brain maturation increases GnRH secretion.
• Decreased sensitivity of gonadotropin to negative
  feedback.
• During late puberty, pulsatile secretion of LH
  and FSH increase during sleep.
• Stimulate a rise in sex steroid secretion.

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Menarche
A critical body weight (47.8kg) must be reached
by a girl to achieved menarche.
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More important than total weight is the shift in
body composition to a greater percent fat (from
16 to 23.5).
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Leptin Anti-obesity or Energy Sufficiency Signal

• Leptin is secreted by fat cells and
  Circulating levels of leptin correlate positively
  with fat stores.
• Administration of leptin accelerates the onset of
  puberty in rodents
• Administration of leptin to rodents and humans
  with congenital or acquired leptin deficiency
  restored fertility.
• It also normalized body weight via variety of
  mechanisms including decreased food intake,
  increased energy expenditure and improved insulin
  sensitivity

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Leptin is a Permissive Factor for the Initiation
of Puberty

• Stimulates rise in testosterone and
  estradiol-17b.
• Produce secondary sexual characteristics.
• Age of onset related to the of body fat and
  physical activity in the female
• Leptin secretion from adipocytes may be required
  for puberty.

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Effects of Leptin Replacement on Body Weight in
Three Leptin-deficient Adult Cousins

• Three first-degree cousins from one large Turkish
  pedigree with a congenital absence of leptin
• Initial body weight, 125 kg on average
• Daily leptin injection for ten months
• J. Licinio, M. Ozata, E. Ravussin, et al.

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Patients Before Treatment
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After Ten Months of Treatment with Human
Recombinant Leptin
The cousins have lost nearly half their body
weight, the man attained puberty and the women
started menstruating cyclically
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Exercised Induced Amenorrhea
Reduced body fat Often accompanied by
amenorrhea Increased risk of osteoporosis due to
low gonadotropin/ estrogen secretion
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Leptin Treatment in Hypothalamic Amenorrhea

• These investigators administered recombinant
  human leptin to eight women with hypothalamic
  amenorrhea
• Six additional untreated subjects served as
  historical controls
• Treatment with recombinant leptin increased mean
  luteinizing hormone levels and pulse frequency
  and resulted in ovulation in three women and in
  withdrawal bleeding in two
• Leptin administration may improve reproductive,
  thyroid, and growth hormone function in women
  with hypothalamic amenorrhea

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Supporting Data (Hamm et al . Biol of
Reproduction 2004)

• Leptin deficient mice were obese, had
  significantly lower uterine weights and exhibited
  delayed vaginal opening
• The leptin deficiency was associated with
  impaired folliculogenesis, increased granulosa
  cell apoptosis and increased expression levels of
  Fas/FasL in primary and secondary follicles

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Mutant obese mice

Leptin
Infertile
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Appetite/Feeding Metabolism
LH/FSH Reproduction?
Metabolic Signal
Exercise
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HYPOTHESIS

• The administration of gonadotropin (GTH) will
  normalize folliculogenesis and induce ovulation
  in prepubertal leptin deficient mice

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OBJECTIVES

• To assess the impact of gonadotropin treatment on
  folliculogenesis in leptin deficient mice
• To assess the effect of gonadotropin
  administration on granulosa cell apoptosis and
  follicular atresia in these animals

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Effect of GTH Treatment on Ovulation in Control
and Leptin Deficient Mice

• No. Ovul. No. Eggs
• Control
• Saline 0/8 0
• GTH 5/8 12.4 4.2
• Ob/Ob
• Saline 0/8 0
• GTH 0/8 0

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Sub normal Response to Exogenous Gonadotropin in
Ob/Ob Mice

• Both control and ob/ob immature mice responded to
  PMSG/hCG treatment but the response was
  attenuated in ob/ob mice
• Only control mice ovulated in response to
  PMSG/hCG treatment
• The number of preantral follicles was higher in
  leptin deficient mice than controls and that
  number was reduced by PMSG/hCG treatment

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CONCLUSIONS

• The data suggest that
• Leptin may influence the process of
  folliculogenesis independent of its ability to
  alter gonadotropin secretion
• Quantitative analysis of the different stages of
  follicle development in the leptin deficient
  animal suggests that folliculogenesis is impaired
  beyond the preantral multilaminar follicle stage
• These observations are consistent with the notion
  that leptin has a direct modulatory effect on
  ovarian function.

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Implantation

• A window of implantation with maximal receptivity
  in women occurs around cycle days 2024
  coincident with the peak serum progesterone
  levels
• Locally secreted cytokines from the embryo and/or
  endometrium control the implantation process

Insert fig. 20.45
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Regulation of Endometrial Receptivity and
Implantation

• Our current knowledge of the autocrine and
  paracrine factors that regulates the process of
  endometrial receptivity and subsequent blastocyst
  implantation is limited
• ß3 Integrin and Leukemia inhibitory factors (LIF)
  are temporally expressed in the endometrium
  during the menstrual cycle with a peak expression
  coinciding with the time of implantation

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The Role of Leptin in Endometrial Receptivity and
Implantation

• Withdrawal of leptin infusion in ob/ob female
  mice shortly after fertilization impairs
  implantation .
• A postovulatory increase in serum leptin
  concentration is associated with successful
  implantation
• Conversely, a low expression level of endometrial
  OB-R is associated with infertility in women
• Leptin increases leukemia inhibitory factor
  (LIF), IL-1 (ligand and antagonist) and ?3
  integrin in human endometrial cells

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glands
Gonzalez et al. Hum Reprod Update 2000
Leptin could play a role in the embryo-maternal
crosstalk
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Hypothesis LPAs will block embryo implantation
by inhibiting leptin signaling and reducing the
endometrial levels of key mediators of
implantation
LPAs
LIF/R
Leptin
b3
OB-R
IL-1 system
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Project Objectives

• To determine if leptin peptide receptor
  antagonists (LPrAs) will effectively block leptin
  signaling in the endometrium in vitro
• To determine the effect of LPrAs on Mouse embryo
  implantation and pregnancy in vivo

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Conclusions

• LPrA2 caused a decrease in well known markers of
  endometrial receptivity that included
  interleukin-1 receptor type I and leukemia
  inhibitory factor and its receptor in rabbit
  endometrial cells

Gonzalez RR, Leavis PC A peptide derived from
the human leptin molecule is a potent inhibitor
of the leptin receptor function in rabbit
endometrial cells. Endocrine. 2003, 2185-95

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Intrauterine injections of LPA-1, LPA-2 or OB-R
antibody on d3 of pregnancy impair mouse embryo
implantation
Ramos et al., Endocrinology 2005
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Contraceptive Effects of Vaginal Application of
LPrA2 for Three days
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Summary

• Although vaginal applications of pegylated LPrA2
  dispersed in CONRAD-gel did not prevent pregnancy
  in 100 of cases, the direct vaginal application
  of unconjugated LPrA2 without the gel completely
  prevented pregnancy in 100 of cases
• PK studies form vaginal application of titrated
  PEG-LPrA2 suggest that the unsatisfactory results
  from contraceptive trials after vaginal
  application could be due to the slow release rate
  of the pegylated peptide from the gel thereby
  leading to the delayed absorption of the peptide
  by the vaginal epithelium

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Objectives

• Leptin has significant role in reproduction.
• Leptin is essential for the onset of puberty.
• Leptin directly regulates ovarian responsiveness
  to gonadotropins.
• Leptin directly regulates endometrial receptivity
  to implantation.

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Acknowledgements

• David Mann
• Ruben Rene Gonzalez
• Winston Thompson
• Ganapathy Bhat
• Christine Stah
• Salandre Cherfils
• Daina Ngugi
• Penelope Cipriani