LM 4 and 5 Flashcards
Why are spermatogonia referred to as true stem cells?
They are capable of infinite self-renewal
Why are spermatogonia excellent target cells for biotechnology?
- Can be genetically engineered in lab
* can be propagated and transferred to produce germ cells and genetically altered offspring
Why has it been historically difficult to manipulate spermatogonia in the lab?
- They are embedded in the seminiferous tubules
- Difficult to obtain a pure sample
- Extremely difficult to culture in lab setting
What was the objective of the Brinster and Zimmerman experiment in 1994 published in PNAS?
To establish methods for successful spermatogonial cell transfer
What were the methods of the Brinster and Zimmerman experiment in 1994 published in PNAS?
- Recipient mice were anesthetized, and the testicles were exteriorized and placed under microscope
- Smeiniferous tubules were injected with the cellular suspension of donor cells in the lumen
- Testicles were re-closed and males recovered
- Mice were then monitored to determine results of donor cell incorporation
Describe the recipient male mice in the Brinster and Zimmerman experiment in 1994 published in the PNAS
- W strain male mice that were deficient in spermatogenesis
* They had been treated with Busulfan to induce the deficiency
Describe the donor cells in the Brinster and Zimmerman experiment in 1994 published in the PNAS
*Mixture of testicular cells from normal male mice
Describe the results of the Brinster and Zimmerman experiment in 1994 published in the PNAS
- 25% of injected testes incorporated donor cells
- Spermatogonia established in ST over time
- Remainder of cells in mixture died out
What were the implications of the Brinster and Zimmerman experiment in 1994 published in PNAS
- Can isolate and hold spermatogonia (in cell mixture) in lab for several hours
- Spermatogonia can establish themselves in ST of sterile recipient males
Describe the methods of the experiment published by Brinster and Avarbock in 1994 in PNAS
- Used same methods of donor cell mixture injected into lumen of the ST of sterile W strain male mice
- Recipients were then observed through puberty and mating
Describe the results of the Brinster and Avabock 1994 PNAS
Recipient sterility was overcome by testicular cell transfer
What was the major problem with spermatogonial cell transfer between the years 1994-2002
*ST spermatogonial cell transfer was okay within species, but no cross-species transfers worked
due to species-specific microenvironment
What was the solution to the major problem facing ST spermatogonial cell trasnfer between the years of 1994-2002
- Transfer whole testis pieces under skin of nude mice (immunodeficient)
- Able to maintain species-specific microenvironment due to being immunocompromised
Describe the experiment done by Honaramooz et al. 2002 in Nature
- Graft a pig testis about 0.5-1mm
- Size expanded to 4-8mm after 10 weeks
- A porcine stem cell was extracted from a 27 week graft
- Intracytoplasmic sperm injection (ICSI) used to produce pig embryos from porcine sperm cells in male mice
Describe the experiment done by Snedaker et al. 2004 J Androl
- Kitten testicular fragments were grafted under the back skin of nude mice
- At 54 weeks, 25% of the grafts had full formed cat spermatozoa
Describe the experiment of Honaramooz et al. 2004 in Bio repro
- Immature Rhesus monkey testicular fragments were grafted under the back skin of nude mice
- ICSI was performed in monkey ova using sperm from the grafts
- Both morulas and blastocysts were produced
What was the major implication of the various testicular fragment grafting experiments done in 2002-2004?
Sperm cells and offspring from one species can be produced in a different species (namely nude mice)
What potential applications arose after the various testicular fragment grafting experiments done in 2002-2004?
- Zoological species propagation
* Treatments for rescue of fertility after cancer treatment in humans
What limitations were still present after the various testicular fragment grafting experiments done in 2002-2004?
- Donor: immature testicular tissue only could be used
* Recipient mice: Must be immunocompromised
Describe the experiment done by Honaramooz et al. in 2003 published in Bio Repro
- AAT transgenic goats used that have an α-1 antitrypsin deficiency
- Testes were removed and testicular cells were prepared
- Cell preps were injected into the rete testis of 5 immature recipient male goats
- The recipients were mated and transgenic offspring were produced
Describe the implications of the experiment done by Honaramooz et al. in 2003 published in Bio Repor
- Speramtogonial cell transfer can be performed in domestic animal species
- Immunocompromised recipients not required
- Simple rete testis infusion technique useable
- Offspring from donor’s spermatogonia produced
- Novel, simple method for making transgenic animals
- A recipient male can produce offspring from another, unrelated site
Describe the experiment of spermatogonial transfer in fish
- Trout spermatogonia was injected into sterile (triploid) salmon fry
- It was incorporated into the gonads and differentiated into spermatogonia and oogonia
- The salmon produced trout offspring
What was the major implication of the experiment involving spermatogonial transfer in fish?
It can be used as a method of species expansion/preservation such as using the blue fin tuna as a target and salmon as the recipient
Why is the dairy industry interested in sexed semen?
- Production of heifers preferred because primary income is fluid milk sales
- Value of the bull is minimal and replacement heifers are always needed
