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N a t a l i a F l ó r e z S a l a z a r
N a t a l i a F l ó r e z S a l a z a r
T h i r d S e m e s t e r
T h i r d S e m e s t e r
B o l i v a r i a n P o n t i f i c i a l U n i v e r s i t y
B o l i v a r i a n P o n t i f i c i a l U n i v e r s i t y
D i a b e t e s
D i a b e t e s
M e l l i t u s
M e l l i t u s
Introduction
Introduction


A t r i a l
A t r i a l
F i b r i l l a t i o n
F i b r i l l a t i o n
______________________________________________________
It is a group of
It is a group of
diseases that results
diseases that results
in too much sugar in
in too much sugar in
the blood (high
the blood (high
blood glucose).
blood glucose).
Irregular and occasionally
Irregular and occasionally
rapid heart rate that often
rapid heart rate that often
causes poor blood supply.
causes poor blood supply.
T h e M A P K 1 0 g e n e i s a c r i t i c a l
T h e M A P K 1 0 g e n e i s a c r i t i c a l
r e g u l a t o r o f D M a s s o c i a t e d A F a n d
r e g u l a t o r o f D M a s s o c i a t e d A F a n d
m a y b e a p o t e n t i a l t h e r a p e u t i c
m a y b e a p o t e n t i a l t h e r a p e u t i c
a p p r o a c h t o t r e a t D M r e l a t e d A F .
a p p r o a c h t o t r e a t D M r e l a t e d A F .
Gen MAPK10
Gen MAPK10
General objective
General objective
U s i n g t i m e - s e r i e s R N A s e q u e n c e
U s i n g t i m e - s e r i e s R N A s e q u e n c e
a n a l y s i s , M A P K 1 0 i s i d e n t i f i e d a s a
a n a l y s i s , M A P K 1 0 i s i d e n t i f i e d a s a
c r i t i c a l g e n e i n d i a b e t e s m e l l i t u s -
c r i t i c a l g e n e i n d i a b e t e s m e l l i t u s -
i n d u c e d a t r i a l f i b r i l l a t i o n i n m i c e .
i n d u c e d a t r i a l f i b r i l l a t i o n i n m i c e .
Materials and methods
Materials and methods
V a r i a n t o f t h e p o l y m e r a s e c h a i n r e a c t i o n .
V a r i a n t o f t h e p o l y m e r a s e c h a i n r e a c t i o n .
U s e d t o a m p l i f y a n d s i m u l t a n e o u s l y q u a n t i f y t h e
U s e d t o a m p l i f y a n d s i m u l t a n e o u s l y q u a n t i f y t h e
D N A a m p l i f i c a t i o n p r o d u c t .
D N A a m p l i f i c a t i o n p r o d u c t .
I t w a s u s e d t o e x t r a c t t o t a l R N A f r o m t h e l e f t
I t w a s u s e d t o e x t r a c t t o t a l R N A f r o m t h e l e f t
a t r i u m a n d r e v e r s e t r a n s c r i b e d i t .
a t r i u m a n d r e v e r s e t r a n s c r i b e d i t .
P r o t e i n s w e r e e x t r a c t e d f r o m c a r d i a c t i s s u e s a n d
P r o t e i n s w e r e e x t r a c t e d f r o m c a r d i a c t i s s u e s a n d
m e m b r a n e s w e r e i n c u b a t e d w i t h a n t i b o d i e s .
m e m b r a n e s w e r e i n c u b a t e d w i t h a n t i b o d i e s .
T h e p u r p o s e o f t h i s t e s t i s t o i d e n t i f y p r o t e i n s
T h e p u r p o s e o f t h i s t e s t i s t o i d e n t i f y p r o t e i n s
b a s e d o n t h e a n t i g e n - a n t i b o d y r e a c t i o n .
b a s e d o n t h e a n t i g e n - a n t i b o d y r e a c t i o n .
R e a l - t i m e P C R A n a l y s i s
R e a l - t i m e P C R A n a l y s i s W e s t e r n B l o t A n a l y s i s
W e s t e r n B l o t A n a l y s i s
______________________________________________________
A s o l u t i o n w a s i n j e c t e d i n t o t h e t a i l v e i n o f
A s o l u t i o n w a s i n j e c t e d i n t o t h e t a i l v e i n o f
m i c e a t 1 0 w e e k s .
m i c e a t 1 0 w e e k s .
I t i s u s e d i n g e n e t h e r a p y p r o t o c o l s , s i n c e t h e y
I t i s u s e d i n g e n e t h e r a p y p r o t o c o l s , s i n c e t h e y
h a v e c h a r a c t e r i s t i c s t h a t m a k e t h e m t o o l s f o r
h a v e c h a r a c t e r i s t i c s t h a t m a k e t h e m t o o l s f o r
t h e t r e a t m e n t o f g e n e t i c d i s e a s e s .
t h e t r e a t m e n t o f g e n e t i c d i s e a s e s .
T h e a p o p t o s i s o f c a r d i o m y o c y t e s i n h e a r t
T h e a p o p t o s i s o f c a r d i o m y o c y t e s i n h e a r t
s e c t i o n s w a s e v a l u a t e d u s i n g t h i s t e s t .
s e c t i o n s w a s e v a l u a t e d u s i n g t h i s t e s t .
T h e a s s a y i s b a s e d o n t h e p r e s e n c e o f b r e a k s i n
T h e a s s a y i s b a s e d o n t h e p r e s e n c e o f b r e a k s i n
t h e D N A w h i c h c a n b e i d e n t i f i e d b y t h e T d T
t h e D N A w h i c h c a n b e i d e n t i f i e d b y t h e T d T
t e r m i n a l .
t e r m i n a l .
I n j e c t i o n o f R e c o m b i n a n t
I n j e c t i o n o f R e c o m b i n a n t
A d e n o - A s s o c i a t e d V i r u s
A d e n o - A s s o c i a t e d V i r u s
T u n n e l A s s a y
T u n n e l A s s a y
______________________________________________________
Results
Results
______________________________________________________
MAPK10 (involved gene), TNF α (cytokine), and tubulin (test
MAPK10 (involved gene), TNF α (cytokine), and tubulin (test
control) are associated with WT and db/db mice followed by
control) are associated with WT and db/db mice followed by
follow-up at week 12, 14, and 16.
follow-up at week 12, 14, and 16.
This image shows an experimental model where a protein is
This image shows an experimental model where a protein is
measured and as time passes, its expression increases and,
measured and as time passes, its expression increases and,
likewise, the damage of the material is evaluated.
likewise, the damage of the material is evaluated.
______________________________________________________
In this graph they show us an immunoassay where the main
In this graph they show us an immunoassay where the main
focus is on showing how in the case of Kv4.3 it is intensified in
focus is on showing how in the case of Kv4.3 it is intensified in
non-mutated mice, Kv4.2 is decreased in mutated mice.
non-mutated mice, Kv4.2 is decreased in mutated mice.
ITNFα is intensified in the db/db mutated mice, as is the
ITNFα is intensified in the db/db mutated mice, as is the
MAPK10 gene and tubulin. It can also be seen how p-stat3,
MAPK10 gene and tubulin. It can also be seen how p-stat3,
which is a mediator of the signaling process, increases
which is a mediator of the signaling process, increases
equally in db/db mice.
equally in db/db mice.
______________________________________________________
______________________________________________________
Discussion
Discussion
______________________________________________________
______________________________________________________
__________________________________________________
__________________________________________________
Author
Author What did he say
What did he say Agree or not
Agree or not
Wang, et al.
Wang, et al.
Liu, et al.
Liu, et al.
Heijman, et al.
Heijman, et al.
Increased AF susceptibility in
Increased AF susceptibility in
db/db mice occurred in
db/db mice occurred in
association with atrial electrical
association with atrial electrical
and estructural remodeling.
and estructural remodeling.
A previous study showed that
A previous study showed that
hyperglycemia contributes to atrial
hyperglycemia contributes to atrial
interstitial fibrosis in an animal
interstitial fibrosis in an animal
model.
model.
Increased atrial fibrosis slows atrial
Increased atrial fibrosis slows atrial
conduction and leads to
conduction and leads to
asynchronous or disorganized
asynchronous or disorganized
conduction.
conduction.
Conclusions
Conclusions
M e d i c a l s t u d i e s ( s u c h a s t h i s
M e d i c a l s t u d i e s ( s u c h a s t h i s
o n e ) w o u l d n o t b e p o s s i b l e
o n e ) w o u l d n o t b e p o s s i b l e
w i t h o u t t h e g r a p h i c s a n d
w i t h o u t t h e g r a p h i c s a n d
a p p l i c a t i o n s t h a t a r e s h o w n
a p p l i c a t i o n s t h a t a r e s h o w n
c o r r e s p o n d i n g t o s t u d i e s o f
c o r r e s p o n d i n g t o s t u d i e s o f
i m m u n o a s s a y s .
i m m u n o a s s a y s .
______________________________________________________
M o l e c u l a r b i o l o g y p r o v i d e s v e r y
M o l e c u l a r b i o l o g y p r o v i d e s v e r y
i m p o r t a n t s t u d i e s f o r t h e
i m p o r t a n t s t u d i e s f o r t h e
a d v a n c e m e n t o f m e d i c i n e a n d i t s
a d v a n c e m e n t o f m e d i c i n e a n d i t s
r e s e a r c h .
r e s e a r c h .
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Diapositivas seminario biologia molecular .pdf

  • 1. N a t a l i a F l ó r e z S a l a z a r N a t a l i a F l ó r e z S a l a z a r T h i r d S e m e s t e r T h i r d S e m e s t e r B o l i v a r i a n P o n t i f i c i a l U n i v e r s i t y B o l i v a r i a n P o n t i f i c i a l U n i v e r s i t y
  • 2. D i a b e t e s D i a b e t e s M e l l i t u s M e l l i t u s Introduction Introduction A t r i a l A t r i a l F i b r i l l a t i o n F i b r i l l a t i o n ______________________________________________________ It is a group of It is a group of diseases that results diseases that results in too much sugar in in too much sugar in the blood (high the blood (high blood glucose). blood glucose). Irregular and occasionally Irregular and occasionally rapid heart rate that often rapid heart rate that often causes poor blood supply. causes poor blood supply.
  • 3. T h e M A P K 1 0 g e n e i s a c r i t i c a l T h e M A P K 1 0 g e n e i s a c r i t i c a l r e g u l a t o r o f D M a s s o c i a t e d A F a n d r e g u l a t o r o f D M a s s o c i a t e d A F a n d m a y b e a p o t e n t i a l t h e r a p e u t i c m a y b e a p o t e n t i a l t h e r a p e u t i c a p p r o a c h t o t r e a t D M r e l a t e d A F . a p p r o a c h t o t r e a t D M r e l a t e d A F . Gen MAPK10 Gen MAPK10
  • 4. General objective General objective U s i n g t i m e - s e r i e s R N A s e q u e n c e U s i n g t i m e - s e r i e s R N A s e q u e n c e a n a l y s i s , M A P K 1 0 i s i d e n t i f i e d a s a a n a l y s i s , M A P K 1 0 i s i d e n t i f i e d a s a c r i t i c a l g e n e i n d i a b e t e s m e l l i t u s - c r i t i c a l g e n e i n d i a b e t e s m e l l i t u s - i n d u c e d a t r i a l f i b r i l l a t i o n i n m i c e . i n d u c e d a t r i a l f i b r i l l a t i o n i n m i c e .
  • 5. Materials and methods Materials and methods V a r i a n t o f t h e p o l y m e r a s e c h a i n r e a c t i o n . V a r i a n t o f t h e p o l y m e r a s e c h a i n r e a c t i o n . U s e d t o a m p l i f y a n d s i m u l t a n e o u s l y q u a n t i f y t h e U s e d t o a m p l i f y a n d s i m u l t a n e o u s l y q u a n t i f y t h e D N A a m p l i f i c a t i o n p r o d u c t . D N A a m p l i f i c a t i o n p r o d u c t . I t w a s u s e d t o e x t r a c t t o t a l R N A f r o m t h e l e f t I t w a s u s e d t o e x t r a c t t o t a l R N A f r o m t h e l e f t a t r i u m a n d r e v e r s e t r a n s c r i b e d i t . a t r i u m a n d r e v e r s e t r a n s c r i b e d i t . P r o t e i n s w e r e e x t r a c t e d f r o m c a r d i a c t i s s u e s a n d P r o t e i n s w e r e e x t r a c t e d f r o m c a r d i a c t i s s u e s a n d m e m b r a n e s w e r e i n c u b a t e d w i t h a n t i b o d i e s . m e m b r a n e s w e r e i n c u b a t e d w i t h a n t i b o d i e s . T h e p u r p o s e o f t h i s t e s t i s t o i d e n t i f y p r o t e i n s T h e p u r p o s e o f t h i s t e s t i s t o i d e n t i f y p r o t e i n s b a s e d o n t h e a n t i g e n - a n t i b o d y r e a c t i o n . b a s e d o n t h e a n t i g e n - a n t i b o d y r e a c t i o n . R e a l - t i m e P C R A n a l y s i s R e a l - t i m e P C R A n a l y s i s W e s t e r n B l o t A n a l y s i s W e s t e r n B l o t A n a l y s i s ______________________________________________________
  • 6. A s o l u t i o n w a s i n j e c t e d i n t o t h e t a i l v e i n o f A s o l u t i o n w a s i n j e c t e d i n t o t h e t a i l v e i n o f m i c e a t 1 0 w e e k s . m i c e a t 1 0 w e e k s . I t i s u s e d i n g e n e t h e r a p y p r o t o c o l s , s i n c e t h e y I t i s u s e d i n g e n e t h e r a p y p r o t o c o l s , s i n c e t h e y h a v e c h a r a c t e r i s t i c s t h a t m a k e t h e m t o o l s f o r h a v e c h a r a c t e r i s t i c s t h a t m a k e t h e m t o o l s f o r t h e t r e a t m e n t o f g e n e t i c d i s e a s e s . t h e t r e a t m e n t o f g e n e t i c d i s e a s e s . T h e a p o p t o s i s o f c a r d i o m y o c y t e s i n h e a r t T h e a p o p t o s i s o f c a r d i o m y o c y t e s i n h e a r t s e c t i o n s w a s e v a l u a t e d u s i n g t h i s t e s t . s e c t i o n s w a s e v a l u a t e d u s i n g t h i s t e s t . T h e a s s a y i s b a s e d o n t h e p r e s e n c e o f b r e a k s i n T h e a s s a y i s b a s e d o n t h e p r e s e n c e o f b r e a k s i n t h e D N A w h i c h c a n b e i d e n t i f i e d b y t h e T d T t h e D N A w h i c h c a n b e i d e n t i f i e d b y t h e T d T t e r m i n a l . t e r m i n a l . I n j e c t i o n o f R e c o m b i n a n t I n j e c t i o n o f R e c o m b i n a n t A d e n o - A s s o c i a t e d V i r u s A d e n o - A s s o c i a t e d V i r u s T u n n e l A s s a y T u n n e l A s s a y ______________________________________________________
  • 7. Results Results ______________________________________________________ MAPK10 (involved gene), TNF α (cytokine), and tubulin (test MAPK10 (involved gene), TNF α (cytokine), and tubulin (test control) are associated with WT and db/db mice followed by control) are associated with WT and db/db mice followed by follow-up at week 12, 14, and 16. follow-up at week 12, 14, and 16. This image shows an experimental model where a protein is This image shows an experimental model where a protein is measured and as time passes, its expression increases and, measured and as time passes, its expression increases and, likewise, the damage of the material is evaluated. likewise, the damage of the material is evaluated.
  • 8. ______________________________________________________ In this graph they show us an immunoassay where the main In this graph they show us an immunoassay where the main focus is on showing how in the case of Kv4.3 it is intensified in focus is on showing how in the case of Kv4.3 it is intensified in non-mutated mice, Kv4.2 is decreased in mutated mice. non-mutated mice, Kv4.2 is decreased in mutated mice. ITNFα is intensified in the db/db mutated mice, as is the ITNFα is intensified in the db/db mutated mice, as is the MAPK10 gene and tubulin. It can also be seen how p-stat3, MAPK10 gene and tubulin. It can also be seen how p-stat3, which is a mediator of the signaling process, increases which is a mediator of the signaling process, increases equally in db/db mice. equally in db/db mice.
  • 9. ______________________________________________________ ______________________________________________________ Discussion Discussion ______________________________________________________ ______________________________________________________ __________________________________________________ __________________________________________________ Author Author What did he say What did he say Agree or not Agree or not Wang, et al. Wang, et al. Liu, et al. Liu, et al. Heijman, et al. Heijman, et al. Increased AF susceptibility in Increased AF susceptibility in db/db mice occurred in db/db mice occurred in association with atrial electrical association with atrial electrical and estructural remodeling. and estructural remodeling. A previous study showed that A previous study showed that hyperglycemia contributes to atrial hyperglycemia contributes to atrial interstitial fibrosis in an animal interstitial fibrosis in an animal model. model. Increased atrial fibrosis slows atrial Increased atrial fibrosis slows atrial conduction and leads to conduction and leads to asynchronous or disorganized asynchronous or disorganized conduction. conduction.
  • 10. Conclusions Conclusions M e d i c a l s t u d i e s ( s u c h a s t h i s M e d i c a l s t u d i e s ( s u c h a s t h i s o n e ) w o u l d n o t b e p o s s i b l e o n e ) w o u l d n o t b e p o s s i b l e w i t h o u t t h e g r a p h i c s a n d w i t h o u t t h e g r a p h i c s a n d a p p l i c a t i o n s t h a t a r e s h o w n a p p l i c a t i o n s t h a t a r e s h o w n c o r r e s p o n d i n g t o s t u d i e s o f c o r r e s p o n d i n g t o s t u d i e s o f i m m u n o a s s a y s . i m m u n o a s s a y s . ______________________________________________________ M o l e c u l a r b i o l o g y p r o v i d e s v e r y M o l e c u l a r b i o l o g y p r o v i d e s v e r y i m p o r t a n t s t u d i e s f o r t h e i m p o r t a n t s t u d i e s f o r t h e a d v a n c e m e n t o f m e d i c i n e a n d i t s a d v a n c e m e n t o f m e d i c i n e a n d i t s r e s e a r c h . r e s e a r c h .