Storage Diseases and Derivation of Mutant StemCell   Lines by Preimplantation      Genetic Diagnosis. Gheona Altarescu, Ra...
INTRUDUCTIONPreimplantation genetic diagnosis( PGD) is a great tool foravoiding the transmission for          genetic dise...
Stem cells• Undifferentiated cell capable of self-  renewal and differentiate into more  specialized cells.
Stem cells             • Self-renewal: the               ability to go through               numerous cycles of           ...
Types of stem cells1. Embryonic stem cells, which are isolated from   the inner cell mass of blastocysts2. Adult stem cell...
Lysosomes                                  Intracellular                                   digestion of     Are membrane  ...
lysosomes                                 ClassificationOver 60 lysosomal enzymes        • Primary: those thatare known. T...
LSD• Lysosomal storage diseases are genetic  disorders in which a genetic mutation affects the  activity of one or more of...
PGDPreimplantation       genetictesting is a technique usedto identify genetic defectsin     embryos       createdthrough ...
Families    Transmission       or not            Diseases of                         lysosomes                            ...
Objective• Present the strategy and outcome of PGD for  four lysosomal storage disorders (  TSD,GD,FD,HS) with the purpose...
Materiales y Métodos• Tay Sachs                       4 (PGD)-5(prenatal diagnosis)                                     Do...
• Gaucher   Familia 2 : heterocigoto /mutación   paterna   Familia 3:mujer GS/marido mutación   84GG -50%
• Fabry :2 parejas : hombre enfermo de fabry                azoospermia no obstructiva en una pareja                hombre...
IVF- estimulación ovárica, recuperación delovocito , fertilización y biopsia                                   congelados-...
Análisis Molecular• Se extrajo el ADN de las células de sangre periférica (  parejas, niños afectados y familiares de prim...
Derivación de líneas y mantenimiento• Derivación y mantenimiento de indiferenciados  Shaare Zedek (SZ) Hunter y células de...
Tabla 3                                      Tasa de                                      embarazo:                       ...
Tabla 3
Tabla 4• De los 28 embriones,  se obtuvieron dos  líneas  de    células  madre    embrionarias  humanas (HESC).• Una de un...
Figura 1
Figura 1 (A)Electroforesis: método delaboratorio en el que seutiliza    una     corrienteeléctrica controlada con lafinali...
Figura 1 (b)• Análisis cromosómico  por     tinción   de  Giemsa, llevado a  cabo en metafase.• Mostró un cariotipo  norma...
Figura 1 (c)               • Se observaron colonias de                 células madre embrionarias                 humanas ...
Discussion                                                   AGREE OR  AUTHOR                  OPINION                    ...
Discussion                                                   AGREE OR   AUTOR                  OPINION                    ...
Conclusions
Conclusions
Concept map
MAPAPAULA
GRACIAS
Prevention of Lysosomal Storage Diseases and Derivation of Mutant StemCell Lines by Preimplantation Genetic Diagnosis.
Prevention of Lysosomal Storage Diseases and Derivation of Mutant StemCell Lines by Preimplantation Genetic Diagnosis.
Prevention of Lysosomal Storage Diseases and Derivation of Mutant StemCell Lines by Preimplantation Genetic Diagnosis.
Upcoming SlideShare
Loading in …5
×

Prevention of Lysosomal Storage Diseases and Derivation of Mutant StemCell Lines by Preimplantation Genetic Diagnosis.

641 views
537 views

Published on

0 Comments
0 Likes
Statistics
Notes
  • Be the first to comment

  • Be the first to like this

No Downloads
Views
Total views
641
On SlideShare
0
From Embeds
0
Number of Embeds
0
Actions
Shares
0
Downloads
3
Comments
0
Likes
0
Embeds 0
No embeds

No notes for slide

Prevention of Lysosomal Storage Diseases and Derivation of Mutant StemCell Lines by Preimplantation Genetic Diagnosis.

  1. 1. Storage Diseases and Derivation of Mutant StemCell Lines by Preimplantation Genetic Diagnosis. Gheona Altarescu, Rachel Beeri, Rachel Eiges, Silvina Epsztejn-Litman, Talia Eldar-Geva, Deborah Elstein, Ari Zimran, Ehud J.Margalioth, Ephrat Levy-Lahad, and Paul Renbaum.Tatiana Gil FrancoPaula E MontoyaMedicine studentsMolecular Biology
  2. 2. INTRUDUCTIONPreimplantation genetic diagnosis( PGD) is a great tool foravoiding the transmission for genetic diseases todescendants. This is a challenging method because haveanalyze de disorder in a single cell , and have to buildprotocols for each specific mutation.Is necessarily that give the results in a short time, that canused in testing two or more indications at once andaccuracy rates approaching 100%; to get this, useinformation of genomic DNA from family and polar bodies ,how give maternal autosomal dominant.
  3. 3. Stem cells• Undifferentiated cell capable of self- renewal and differentiate into more specialized cells.
  4. 4. Stem cells • Self-renewal: the ability to go through numerous cycles of cell division while maintaining the undifferentiated state. • Potency: the capacity to
  5. 5. Types of stem cells1. Embryonic stem cells, which are isolated from the inner cell mass of blastocysts2. Adult stem cells, which are found in various tissues.
  6. 6. Lysosomes Intracellular digestion of Are membrane Hydrolytic macromolecules, cytoplasmic enzymes old cell organelles parts, and microorganisms
  7. 7. lysosomes ClassificationOver 60 lysosomal enzymes • Primary: those thatare known. There is a contain enzymes solon.hydrolyse for each type of • Secondary: besidesbiological molecule: containing enzymes, also• Peptidases – hydrolyse digestion materials. proteins• DNAases – hydrolyse DNA• RNAases – hydrolyse RNA• Lipases – hydrolyse lipids• Phosphatases – hydrolyse phosphates• Glucosidases – hydrolyse glycogen
  8. 8. LSD• Lysosomal storage diseases are genetic disorders in which a genetic mutation affects the activity of one or more of the acid hydrolases. In such diseases, the normal metabolism of specific macromolecules is blocked and the macromolecules accumulate inside the lysosomes, causing severe physiological damage or deformity.
  9. 9. PGDPreimplantation genetictesting is a technique usedto identify genetic defectsin embryos createdthrough in vitro fertilization(IVF) before pregnancy.Preimplantationgenetic diagnosis (PGD)refers specifically to whenone or both geneticparents has a knowngenetic abnormality andtesting is performed on an
  10. 10. Families Transmission or not Diseases of lysosomes LSDStem cells (TSD,GD,FDHS) PGD
  11. 11. Objective• Present the strategy and outcome of PGD for four lysosomal storage disorders ( TSD,GD,FD,HS) with the purpose of avoiding the transmission for genetic diseases and termination of pregnancy in couples at risk of transmitting of these disorders.
  12. 12. Materiales y Métodos• Tay Sachs 4 (PGD)-5(prenatal diagnosis) Double carrier
  13. 13. • Gaucher Familia 2 : heterocigoto /mutación paterna Familia 3:mujer GS/marido mutación 84GG -50%
  14. 14. • Fabry :2 parejas : hombre enfermo de fabry azoospermia no obstructiva en una pareja hombres serán sanos –mujeres portadoras(ligado al X)• Síndrome de no implantar portadores Hunter1 y 2 : hermanas –CVS3: mujer –análisis prenatal –interrupción –
  15. 15. IVF- estimulación ovárica, recuperación delovocito , fertilización y biopsia congelados- descongelados: Valerato de estradiol oral (Estrofem 4- 8mg al día) y vaginal Utrogestan (progesterona micronizada 900 mg / día). Cuerpo polar y blastómero
  16. 16. Análisis Molecular• Se extrajo el ADN de las células de sangre periférica ( parejas, niños afectados y familiares de primer grado ).• Para cada enfermedad:Marcadores polimórficos de microsatélites que rodean el gen enfermo se identificaron y los marcadores informativos usados, se crearon haplotipos para cada familia. Estos marcadores y las mutaciones familiares se usaron para el desarrollo de ensayos múltiples de una sola célula.Una reacción de PCR múltiple se utiliza .Sólo las muestras que fueron informativas para un mínimo de tres marcadores polimórficos se consideraron para el diagnóstico.
  17. 17. Derivación de líneas y mantenimiento• Derivación y mantenimiento de indiferenciados Shaare Zedek (SZ) Hunter y células de Gaucher se llevaron a cabo de acuerdo con protocolos aplican rutinariamente en blastocitos diagnosticados como mutante
  18. 18. Tabla 3 Tasa de embarazo: Análisis de 38% 329 56 ciclos ovocitos de PGD20Familiasconmutaciones
  19. 19. Tabla 3
  20. 20. Tabla 4• De los 28 embriones, se obtuvieron dos líneas de células madre embrionarias humanas (HESC).• Una de un embrión mutante Hunter hembra.• Otra de compuestos heterocigotos 84GG/N370S para la
  21. 21. Figura 1
  22. 22. Figura 1 (A)Electroforesis: método delaboratorio en el que seutiliza una corrienteeléctrica controlada con lafinalidad de separarbiomoleculas según sutamaño y carga eléctrica através de una matrizgelatinosa. (1937) Estas nuevas líneas presentan características típicas de células madre embrionarias humanas, que expresan un panel de marcadores no diferenciados, como NANOG, Oct4, Sox2, y REX
  23. 23. Figura 1 (b)• Análisis cromosómico por tinción de Giemsa, llevado a cabo en metafase.• Mostró un cariotipo normal, 46XX humano para la línea celular Hunter y 46XY para la línea de
  24. 24. Figura 1 (c) • Se observaron colonias de células madre embrionarias humanas (HESC) y cuerpos embrioides. • HESC: células que poseen la capacidad de dividirse por largos periodos, se pueden diferenciar en las células de distintos linajes. (Pluripotentenciales). • Cuerpos embrioides: son agregaciones espontáneas de HESC que se
  25. 25. Discussion AGREE OR AUTHOR OPINION DISAGREE In the second couple there was no known infertility but since more than half of female carriers of Fabry K. Toyooka disease develop symptoms during life [22], they preferred medical sex election for males. Since PGD was firstG. L. Harton, M. performed in 1991, by using different strategies, the De Rycke, F. technique has become very
  26. 26. Discussion AGREE OR AUTOR OPINION DISAGREEP. Kozlowski, A. Both of these invasive methods are accompaniedKnippel, and R. by a small risk of abortion Stressig. due to the procedure [28]. One Gaucher stem cell line was derived from mutant A. Ribner, G. embryos caring 84GG and Altarescu, A. N370S mutations is of Zimran, and particular of interest due to recent evidence of D. Elstein. correlations between Parkinson disease and
  27. 27. Conclusions
  28. 28. Conclusions
  29. 29. Concept map
  30. 30. MAPAPAULA
  31. 31. GRACIAS

×