MALE INFERTILITY
Development of in vitro spermatogenesis
and genetic analysis
Huleihel Mahmoud, Ph.D
The Shraga Segal Department of Microbiology, Immunology and Genetics, Faculty of Health Sciences, Ben-Gurion University of the Negev
InfertilityInfertility
* Infertility affects 1/5 couples of reproductive age and in half male infertility is the sole or a contributory cause.
*Testicular disorders affecting spermatogenesis account for 2/3 of cases and the majority are of unknown etiology.
* Genetic and epigenetic factors are involved in human infertility. Appromimately10% are defined.
Spermatogenic Dysfunction
Possible Etiologies:
1. Congenital (inherited, de novo, etc.)
2. Acquired (infection, environmental factors, etc.)
3. Iatrogenic (chemotherapy, radiotherapy, etc.)
There has been a gradual decrease in male fertility world wide
Goals
We propose to address testicular dysfunction by:
1. Identifying specific genes that can contribute to spermatogenic dysfunction.
2. Developing new in vivo and in vitro technologies that enable propagation and differentiation of spermatogonial stem cells (SSCs) into functional sperm.
Small colony Medium colony Large colony
ACB
Murine colony formation in agar4 weeks culture
Development of murine sperm-like cells in agar
Small Medium Large
Colony formation in agar/methylcellulose
Non human primate (NHP) system
GFR- SALL-4 VASA
CREM-1NC
Expression of spermatogenesis markers in juvenile NHP testicular germ cells cultured in methylcellulose
ACROSIN
Growth of prepubertal (7 y) testicular cells (germ cells) in culture(2 weeks)
Immunofluorescence staining of spermatogenesis markers from prepubertal boys before and after 5 weeks of culture
Pre-meiotic (SSC)Protamine
Post-meioticMeioticLDH
CD-9GFR-C-Kit
Pre-meiotic markers (SSC)Before
After
CD-9 GFR-
Development of colonies from cells isolated from biopsies of adult azoospermic patients
Pre-meiotic
(SSC)
Meiotic Post-meiotic
Weeks C-kit GFR- CD9 CREM LDH Protamine Acrosin
Before
culture0 - + - - - - -
After
culture5 - + _ - + - +
RNA expression levels of spermatogenic markers in colonies developed in methylcellulose from cells isolated from
biopsies of adult azoospermic patients
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