This ppt will give you all information about various animal models for screening of drugs that are useful in the treatment of heart failure. This ppt is made by various authenticated research articles and books. ppt include- In-vivo models: 1. Rat models 2. Dog models 3. Rabbit models 4. Guinea pig models 5. Syrian hamster models 6. Murine models 7. Zebrafish models 8. Swine models 9. Transgenic mice 10. Other models (22 other models) In-vitro models: 1. Isolated hamster cardiomyopathic heart 2. Isolated cat papillary muscle 3. Ouabain binding YouTube Channel Link for drug information: https://www.youtube.com/channel/UC49iKtopfbsV3ggHy7pmAug/about

1. Animal models for
screening agents useful in
heart failure
Presented by: Akash Agnihotri

2. Introduction- Heart Failure (HF)
Heart Failure
• Heart can’t supply enough blood to meet the body’s demand
Heart Failure
Systolic HF Diastolic HF
• Ventricles can’t pump
blood hard enough
• Can't contract completely
• Ventricles not enough
filled with blood
HFrEF HFpEF

3. Introduction- Heart Failure (HF)
• HF- leading cause of morbidity and mortality

4. Risk factors for HF:
• Heart failure with reduced ejection
fraction (HFrEF)
• Heart failure with preserve ejection
fraction (HFpEF)
• COPD- Chronic Obstructive
Pulmonary Disease
• LVEF- Left Ventricular Ejection
Fraction

5. Introduction- Heart Failure (HF)
• Despite a number of important therapeutic advances
• prevalence, mortality, and cost associated with HF continue to grow
• Current treatments primarily:
• Slow the progression of this syndrome
• There is a need to develop novel preventative and reparative therapies
• Development of these novel HF therapies requires testing of the
putative therapeutic strategies in appropriate HF animal models

6. Drug Development Pipeline:

7. Stages in
development
of heart
failure &
recommended
therapy by
stage
According to joint American college of cardiology & American Heart Association Guidelines

8. Animal Models for Heart Failure

9. Characteristics of the ideal animal model for
the study of congestion in heart failure

10. Animal Models for Heart Failure
According to joint American college of cardiology & American Heart Association
Guidelines
1. Valvular Lesions That Cause HF
2. Dilated Cardiomyopathy
3. Hypertensive Heart Disease
4. Restrictive Cardiomyopathy
Aortic stenosis
Mitral Regurgitation
Small/Large Animal
Models for AS/MR
Coronary ligation
Coronary Microembolization
Toxic Models Pacing-Induced
Tachycardia
Spontaneously hypertensive rat,
Aortic banding
Rodent models of acquired & hereditary
RCM

11. Animal models for Heart Failure
In vitro models:
1. Isolated hamster
cardiomyopathic heart
2. Isolated cat papillary muscle
3. Ouabain binding
In vivo models:
1. Rat models
2. Dog Models
3. Rabbit models of heart failure
4. Guinea pig model
5. Syrian hamster
6. Genetic model
7. New therapeutic targets in CHF

12. Pros and cons of Animal models

13. In-Vivo Models

14. Animal models for Heart Failure-
In-vivo models
I) Rat models
• Rat coronary ligation model
• Rat aortic banding
• Dahl salt sensitive rats
• Spontaneous hypertensive rat
• Spontaneous hypertensive-
heart failure rats (SH-HF)
II) Dog models
• Chronic rapid pacing
• Volume overload
• Coronary artery
ligation and
microembolization
III) Rabbit models of heart
failure
• Volume and pressure overload
• Tachycardia pacing
• Doxorubicin cardiomyopathy
IV) Guinea pig model
• Cardiac insufficiency
V) Syrian hamster
• Cardiomyopathic hamster
VI. Murine models
• By ischemia
• Pressure overload and
neurohumoral activation
VII. Zebrafish models
VIII. Swine models
XI. Transgenic mice
X. Other Models

15. I. Rat models for Heart Failure

16. I. Rat Models for Heart Failure
1. Rat coronary ligation model
2. Rat aortic banding
3. Dahl salt sensitive rats
4. Spontaneous hypertensive rat
5. Spontaneous hypertensive-heart failure rats (SH-HF)

17. I. Rat Models of Heart Failure
• Relatively inexpensive and because of short gestation period
• A large sample size can be produced in a short period of time
• Extensively used
• Study long-term pharmacological interventions

18. 1. Rat coronary ligation model
Purpose and Rationale
• Incomplete or complete ligation of left coronary artery causes ischemia
of cardiac muscle
• Failure is associated with left ventricular dilatation
• Reduced systolic flow
• Increase in filling pressure

19. 1. Rat coronary ligation model
Procedure
• Male Spargue Dawley rats- anaesthetized with hexobarbitol (200mg/kg )
• Trachea cannulated- artificial respiration provided
• Chest cavity exposed- LAD coronary artery isolated
• Ligature placed and cavity sutured back

20. 1. Rat coronary ligation model
Procedure
• After 4 weeks chest cavity opened- carotid and jugular vein cannulated
• Filling pressure, systolic, diastolic and mean blood pressure are
measured
• Animal sacrificed after hemodynamic parameters tested
• Isolated hearts and used study calcium channels, Sarcoplasmic
Reticulum ATPase (SR-ATPase) and protein levels

21. 1. Rat coronary ligation model
Evaluation
• Progression of left ventricular dysfunction and myocardial failure
• associated with neurohumoral activation(like in human)
• Depressed myocardial function is associated with altered calcium
transients
• The density of L-type calcium channels, SR-Ca2+ - ATPase and protein
levels decrease continuously with increasing severity of congestive
heart failure

22. 1. Rat coronary ligation model
Disadvantage:
• High initial mortality and induction of mild heart failure in most cases
Useful:
• For long term studies of pharmacological interventions on the
neurohormonal activation

23. 2. Rat Aortic Banding
Purpose and Rationale:
• Restriction of blood flow to aorta
• Induces pressure overload - Hypertension and Congestive heart failure
• After several weeks:
• Ventricular ACE activity decrease again to normal values
• May be related to normalization of wall stress with increasing cardiac
hypertrophy
• Furthermore, after several months:
• Animals goes into cardiac failure

24. 2. Rat Aortic Banding
Procedure:
• Sprague–Dawley rats (250–280 g), fasted for 12 h before surgery
• Anesthetized with 200 mg/kg hexobarbitone
• Abdomen is shaved, moistened with a disinfectant and opened by a cut
parallel to the linea alba
• Aorta is isolated
• Ligature placed and cavity sutured back

25. 2. Rat Aortic Banding
Procedure:
• In sham operated controls no banding is performed
• While in the test group animals are administered drugs for 6 weeks
• After 4–6 weeks heart failure develops in these animals

26. 2. Rat Aortic Banding
Evaluation:
• Total cardiac mass, weight of left and right ventricle of treated rats are
compared with sham-operated controls
• Heart failure is associated with increased myosin heavy chain mRNA and
atrial natriuretic factor mRNA
• This model seems to be well suited for studying the transition from
hypertrophy to failure at the level of myocardium

27. 3. Dahl Salt Sensitive Rats
Purpose and Evaluation:
• To study the transition from compensated hypertrophy to failure
• This strain of rats develop systemic hypertension after receiving a high-
salt diet

28. 3. Dahl Salt Sensitive Rats
Procedure:
• Sprague-Dawley rats (250–300 g)
• Drinking water is replaced with 1% NaCl saline solution
• Test drug rats are administered the drug orally for 1 month
• After completion- Both groups sacrificed- Heart observed

29. 3. Dahl Salt Sensitive Rats
Evaluation:
• Total cardiac mass, weight of left and right ventricle are weighed
compared
• It is observed that:
• Sham control group develop concentric left ventricular hypertrophy at 8
weeks
• followed by marked left ventricular dilation and overt clinical heart failure at 15-
20 weeks
• Failing heart dies within a short period of time
• The ability of the test drug to reverse these changes is studied

30. 4. Spontaneous Hypertensive Rat
Purpose and Rationale:
• Well-established model of genetic hypertension
• Cardiac pump function is preserved at 1 year of age
• At 18-24 months, cardiac failure develops which includes reduced
myocardial function and increased fibrosis
• Altered calcium cycling is observed
• An increased number of apoptotic myocytes are observed
• Suggested that apoptosis might be a mechanism involved in reduction of
myocyte mass that accompanies transition from stable compensation to heart
failure

31. 4. Spontaneous Hypertensive Rat
Procedure:
• Animals are divided into two groups.
• Test group animals- drug is administered orally for 1 month
• Sham control group animals- no drug treatment is given
Evaluation:
• Animals- sacrificed- their hearts are processed for:
• Estimation of number of apoptotic cells
• Sarcoplasmic reticulum calcium pump mRNA levels
• Expression of genes encoding for extracellular matrix and results compared

32. 5. Spontaneous Hypertensive-Heart Failure Rats
(SH-HF)
Purpose and Rationale:
• SHR that Develop heart failure before 18 months- selective bred
• Development HF occurs earlier in rats which carry
• facp gene (corpulent gene) that encodes a defective leptin receptor (SH-HF/Mcc-
facp)
• These having increased:
• Plasma renin activity
• ANP (atrial natriuretic peptide)
• Aldosterone levels
These changes might be related to spatial remodeling between L-type calcium
channels and ryanodine receptors

33. 5. Spontaneous Hypertensive-Heart Failure
Rats (SH-HF)
Procedure:
• Animals divided into two groups
• Group 1- test drug group (administered drug for one month, orally)
• Group 2- sham control group (untreated)
• After completion of the experiment comparisons between:
• Plamsa renin activity, ANP (Atrial Natriuretic Peptide)
• Aldosterone levels
• Rynodine receptor density, sarcoplasmic reticulum calcium uptake
• Endothelial nitric oxide synthase (NOS) activity

34. II. Dog Models for Heart Failure

35. II. Dog Models of Heart Failure
• Large animal models, allow to study
• Left ventricular function and volumes more accurately than rodent models
• Furthermore, in dog like human myocardium β-myosin heavy chain
isoform predominates
• Excitation contraction coupling processes seem to be similar to the
human myocardium
• Dog models are costly

36. 1. Chronic Rapid Pacing
Purpose & Rationale:
• Chronic rapid pacing at heart rates above 200 bpm
• In previously healthy dogs within several weeks produces the syndrome
of CHF
Procedure:
• Adult male dog, 18-25kg- anaesthetized with phenobarbital 30mg/kg
• Airway maintained- Chest cavity opened by 3-4cm long
• Thoracotomy- Heart Exposed

37. 1. Chronic Rapid Pacing
Procedure:
• Ventricular pacing lead is attached to apex of heart
• Pacemaker is programmed to pace at 240–260 beats/min for 2–4 weeks
• Cavity closed after placing heart back
• Significant HF develops in 4 weeks
• HF developed for 6 more weeks
• There is bilateral ventricular dilatation over 3-4 weeks
• Test drug is administered by SC and IM injection

38. 1. Chronic Rapid Pacing
Evaluation:
• Ejection fraction decreases- decreased CO and increased resistance
• There is time dependent neurohormonal and hemodynamic
abnormalities
• HF is reversible if pacing is stopped
• 2 groups are compared for parameters like
• Ejection fraction, CO, systemic vascular resistance
• Plasma renin and ANP levels are also compared

39. 2. Volume Overload
Purpose and Rationale
• Prolonged volume overload can lead to HF
• Produced by- creation of an arteriovenous fistula
• Destruction of the mitral valve

40. Types of hemodynamic overload

41. 2. Volume Overload
Procedure
• Dogs (12–15 kg)- anesthetized with pentobarbital (30 mg/kg) IP
• Maintained on artificial respiration (20–24 strokes/min)
• Thoracotomy is performed and heart is exposed
• Chronic experimental mitral regurgitation is produced
• Within 3 months:
• Left ventricular hypertrophy
• Dilation and development of overt clinical heart failure
• Significant heart failure develops by 4 weeks and continues for upto 10 weeks
• Test drugs are administered SC or IM over a period of 14 days

42. 2. Volume Overload
Evaluation
• Neurohumoral activation including local activation of Renin Angiotensin
System (RAS) is observed in CHF dogs
• Test group and sham control group compared
• Used to study the influence of chronic β-adrenoceptors blockade on
myocytes and left ventricular function

43. 3. Coronary Artery ligation and microembolization
• Used to produce myocardial infarction and CHF in dogs
Procedure: (fig on next slide)
• Adult male dog (10-12kg)- anaesthetized with pentobarbitol 30mg/kg
• Airway maintained
• Transducer introduced from femoral artery for peripheral pressure data
• A microtip catheter inserted via carotid for measuring ventricular
pressures
• Heart is exposed- polystearyl microspores are injected through
angiogram catheter- stepwise elevation of LVEDP- target 16-18mmHg

44. 3. Coronary Artery ligation and microembolization
Polystearyl microspores are injected through angiogram catheter
Some microspheres-
• Beads of agarose/polystyrene
• Thrombin and autogenous blood
with fibrinogen intracoronary
injection can be used

45. 3. Coronary Artery ligation and microembolization
Evaluation:
• Recordings are obtained before and after treatment with test drug
Several Disadvantages:
• Time consuming and costly
• Co-lateral circulation- comparison between man and dog is difficult
• High mortality and morbidity (arrythmia)

46. III. Rabbit Models for Heart Failure

47. III. Rabbit Models of Heart Failure
• Less expensive than dog models
• Interesting similarities to human heart
Models:
• Volume and pressure overload
• Tachycardia pacing
• Doxorubicin cardiomyopathy

48. 1. Volume and Pressure Overload
Purpose and Rationale:
• Volume overload, pressure overload or both induced HF
• Chronic severe aortic regurgitation- systolic dysfunction-HF

49. 1. Volume and Pressure Overload
Procedure:
• Rabbits- anaesthetized with pentobarbitone sodium 35mg/kg,IP
• Trachea cannulated to maintain artificial respiration
• Chest cavity opened and aortic insufficiency created by destroying valve
• After 14 days- aortic constriction using PVC clamp
• HF occurs within 4 weeks

50. 1. Volume and Pressure Overload
Evaluation:
• Animal sacrificed after experimental protocol
• HF is associated with alteration in β-adrenoceptors levels
• Protein and mRNA levels of Na+/Ca2+ exchanger increased
• The ability of test drug to reverse these changes is observed
• Mimics alteration of myocardial function
• Used to study- changes in excitation contraction coupling in
hypertrophy an failing agent

51. 2. Tachycardia Pacing
• Chronic rapid pacing- 350–400 beats/min
• Produce myocardial depression, hemodynamic and neurohumoral signs of heart
failure
Procedure:
• Rabbits anaesthetized with pentobarbitone sodium 35mg/kg
• Artificial respiration- Chest cavity opened- pacing lead
• Pace 350-400 beats/min is set
• 4-6 weeks- HF occurs
• 2 groups formed- Test and Sham

52. 2. Tachycardia Pacing
Evaluation:
• Animal sacrificed after experiment
• Heart is weighed
• Parameters-
• Hemodynamic
• Plasma Renin activity
• Weight of hearts compared

53. 3. Doxorubicin Cardiomyopathy Model
Purpose and Rationale:
• Doxorubicin exhibits acute and chronic cardiotoxicity
• Induce HF in various animal species
Mechanism:
• Free radical generation and lipid peroxidation
• Reactive sulphydryl groups
• Binding to channel regulatory sites or
• Inhibition of mRNA and protein synthesis

54. 3. Doxorubicin Cardiomyopathy Model
Procedure:
• Rabbits of both sexes and various strains(5-kg)
• Doxorubicin 1mg/kg IV twice weekly for 6-9 weeks
• Test group- 4-6 weeks IP or SC
• After experiment- Anesthetized with pentobarbitone sodium (35
mg/kg)IP
• Measure left ventricular end diastolic pressure (LVEDP) and dP/dt

55. 3. Doxorubicin Cardiomyopathy Model
Evaluation:
• Heart is processed for immunohistochemical tests
• Chronic doxorubicin causes impairment of cardiac contractility
• Decreased gene expression of Ca+ induced Ca+ release channels in SR-
Rynodine receptor
• C-13 hydroxy metabolite (doxorubicinol), formed in the heart
hypothesized
• Contribute to anthracycline cardiotoxicity
• Ability of test drug to reverse these conditions is observed in both
groups

56. IV. Guinea Pig Model for Heart Failure

57. IV. Guinea Pig Model- Cardiac Insufficiency
Purpose and Rationale
• 8 weeks of cardiac banding of descending thoracic aorta- overt CHF
• Very much similar to human heart failure

58. IV. Guinea Pig Model- Cardiac Insufficiency
Procedure:
• Male guinea pigs, 250-400g- anaesthetized with ether
• Chest cavity opened- heart exposed- aorta located and ligated
• Symptoms of CHF developed- 80% in one day
• Lung weight, relative heart weight are increased

59. IV. Guinea Pig Model- Cardiac Insufficiency
Evaluation:
• Lung and heart weight increases due to failure
• Decrease in SR-Ca2+ ATPase and phospholamban is seen in HF
• Signs and symptoms of HF seen
• Ability of test drug to reverse these signs- observed

60. V. Syrian Hamster Model
for Heart Failure

61. V. Syrian Hamster- Cardiomyopathic Hamster
Procedure:
• Cardiomyopathic strains of the syrian hamsters- widely used
• Autosomal recessive mode of inheritance
• leads to degenerative lesions in all striated muscles (myocardium)
• HF developed- 7-10 months
• Time dependent change in myosin isoform expression-
• Cardiomyopathy
• Fibrosis and Calcium deposition
• Overlapping period of reactive hypertrophy
• Depressed myocardial function

62. V. Syrian Hamster- Cardiomyopathic Hamster
Evaluation:
• Test drugs- administered by SC and IM route for 14 days
• Ability of drug to reverse the condition is observed

63. VI. Murine models for Heart Failure

64. VI. Murine models for Heart Failure
Models of heart failure induced by ischemia
1. Myocardial infarction induced by permanent LAD-ligation
2. Myocardial infarction induced by transient LAD-ligation (ischemia–reperfusion)
Models of heart failure induced by pressure overload and
neurohumoral activation
3. Transverse aortic constriction (TAC)
4. Chronic subjection to angiotensin II
5. Chronic subjection to deoxycorticosterone acetate(DOCA)
6. Model of heart failure induced by viral myocarditis

65. 1. Myocardial infarction induced by permanent
LAD-ligation
• Permanent LAD-ligation in mice results in acute myocardial injury due to
ischemia, which is defined as MI
• In humans, type 1 MI is caused by the disruption of an atherosclerotic
plaque and results in myocyte necrosis, which is often accompanied by
ST-elevations in the ECG (ST-elevated MI, STEMI)
Procedure:
• Adult mice anaesthetized- mechanically ventilated- Heart surgery (incision- 3rd left
intercostal space)
• LAD is permanently ligated with a suture
• Causes ST-elevation- then MI
o Cardiac tissue can be incubated with colourless,
hydrophilic triphenyltetrazolium chloride
immediately after harvesting

66. 2. Myocardial infarction induced by transient
LAD-ligation (ischemia–reperfusion)
• Surgical preparation is performed similar to the procedure of inducing permanent
LAD-ligations
• Ligation is maintained for 20-60 min
• Ligation <30 min may not provoke ischemia-induced myocardial injury
• Ligation of 60-90 min results in irreversible CM death and complete infarction of the
area at risk
• 45–60 min of ischemia is recommended for the transient LAD-ligation

67. 3. Transverse aortic constriction (TAC)
• Performed by constricting the transverse aortic arch between the
brachiocephalic trunk and the left carotid artery
• Mice- anaesthetized- artificially ventilated- chest is entered by an upper
partial sternotomy- suture is tied around aortic arch against a cannula
• Pressure gradient across the stenosis of more
than 40 mmHg
• Non-invasive pulse-wave doppler analysis
can be used to measure this gradient

68. 3. Transverse aortic constriction (TAC)
• 6 weeks after TAC- hypertrophic murine heart, whose ventricular mass
is enlarged by approx. 200%
• Recent studies shows-
Some mice will not proceed
from HFpEF to HFrEF after TAC
at all
• Therefore, it is suggest that-
Examine cardiac function by
means of echocardiograph
prior to further investigations

69. 4. Chronic subjection to angiotensin II
• Osmotic minipumps, which subsequently release Ang II at a defined
rate, are implanted subcutaneously
Procedure:
• Anaesthesia- maintained during surgery
• No artificial ventilation is needed during the procedure(easier)
• Typically, mice are subjected to Ang II for a time frame of 2–8 weeks
Limitations:
• Even at a given infusion rate (1.4 mg/kg per day) & Ang II (8 weeks)
• Cardiac remodelling and dysfunction strongly depends on the specific mouse strain:
• C57BL/6 mice- shows display concentric hypertrophy
• Balb/c mice- shows congestive HF

70. Fig. shows-
Reciprocal interactions
between pressure overload,
activation of the renin-
angiotensin-aldosterone-
system (RAAS), and heart
failure in men and mice

71. 4. Chronic subjection to deoxycorticosterone
acetate (DOCA)
• Chronic subjection to the aldosterone analogue deoxycorticosterone acetate
accompanied by unilateral nephrectomy and high salt diet (DOCA) promotes
hypertension and the subsequent development of HF in mice
Procedure:
• Animals- anaesthetized- unilateral nephrectomy- drinking water that
contains 1% sodium chloride is applied
• Few days after nephrectomy- pellets that release the aldosterone
analogue DOCA at a constant rate are implanted subcutaneously

72. 5. Model of heart failure induced by viral myocarditis
• Coxsackievirus B3 is a non-enveloped, single-stranded, positive-sense
RNA virus within the family of the picornaviredae
• Commonly, 104-107 plaque-forming units of the heart passaged
enterovirus CVB3 are injected IP
• Frequently used one is- Nancy-strain
• Appropriate viral infection can be proven:
• Loss of bodyweight and detection of viremia in small serum samples
• Which typically occurs around day 2-4 post injection (p.i)
• Tissue sampling and hemodynamic assessments performed
• From day 6 to 8 during the acute phase of myocarditis
• Around day 28 during the chronic phase

73. Fig-
The course of viral myocarditis
induced by CVB3 in mice

74. VII. Zebrafish models for Heart Failure

75. VII. Zebrafish heart failure models
• In recent years, the zebrafish, Danio rerio, has emerged as an excellent
genetic and embryonic model system
• Favorable features, including
• Small size, Optical translucency during early development
• Rapid embryonic development, Genetic resemblance to humans and Disease
characteristics similar to humans
A pair of adult zebrafish
Produces hundreds of eggs
Embryos at one-cell stage
24 h post fertilization
72 h post fertilization

76. VII. Zebrafish heart failure models
• The zebrafish embryo can tolerate the absence of blood flow because its
oxygen is delivered by diffusion rather than by the cardiovascular
system, making it an excellent model for studying heart failure
• The cardiac natriuretic peptide genes (nppa and nppb) are known
markers of cardiomyocyte hypertrophy and heart failure
• Models:
1. Sarcomere genes that induce zebrafish models of heart failure
2. Calcium homeostasis associated genes in heart failure
3. Heart failure models induced by mutations mitochondrial associated genes
4. Chemical compounds that trigger heart failure in zebrafish
*To visualize the heart chambers, a transgenic strain of zebrafish larvae that expresses Green fluorescent
protein (GFP) exclusively in the cardiac myosin light chain 2 (cmlc2) [Tg (cmlc2:gfp)] is used

77. Established zebrafish genetic models of heart
failure
HCM:
Hypertrophic
cardiomyopathy
DCM:
Dilated
cardiomyopathy
1.
2.

78. 3. Heart failure models induced by mutations
mitochondrial associated genes
• Mitochondria generate reactive oxygen species (ROS) that mediate the
inotropic and hypertrophic effects of sympathetic and renin-
angiotensin-aldoesteron systems
• Changes in the composition and function of the mitochondrial
proteome
• Tom70, a receptor for translocases in the outer mitochondrial membrane (Tom)
complex, is downregulated in pathological hypertrophic hearts
• Knockdown of Tom70 induces pathological cardiac hypertrophy both in vivo and in
vivo

79. 3. Heart failure models induced by mutations
mitochondrial associated genes
• Tom70 morpholino-injected zebrafish embryos
• Tom70 morphants develop thicken ventricular wall, increased cardiomyocyte
size and reduced ventricular contractility, which are typical features of
pathological cardiac hypertrophy
• Tom70-targeted optic atrophy-1 (Opa1) (defective)-- triggers oxidative stress,
which contributes to pathological cardiomyoctye hypertrophy

80. 4. Zebrafish heart failure models induced by
Chemical

81. 5. Other Zebrafish Model
• Larval zebrafish have recently shown many advantages for human
disease studies and drug discovery
• This model was further validated with 6 FDA-approved heart failure
therapeutic drugs
• LCZ696, Digoxin, Irbesartan, Metoprolol, Enalapril, and Hydrochlorothiazide
• 2 China FDA (CFDA)- approved heart failure therapeutic medicines
• Qiliqiangxin capsule and Shenmai injection
• Also, many heart failure therapeutic drugs- In-vivo screening and efficacy can be
studied by this model

82. Pharmacological impact of common human cardiovascular
drugs on zebrafish

83. VIII. Swine Models for Heart Failure

84. VIII. Swine Models for Heart Failure
• Due to their greater anatomical similarity to humans, and the ability to
make lesions of the same size swine models are used
• Also, arterial anatomy and collateral coronary circulation in pigs can
better mimic those of humans
• In addition, infarct size in porcine models can be precisely predicted
• Among porcine models:
• Balloon occlusion of LAD coronary artery is the most commonly used
model

85. VIII. Swine Models for Heart Failure
1. Myocardial infarction models/Ischemia/HF
• Balloon occlusion of LAD coronary artery
• Microembolization of coronary arterioles (under fluoroscopy)
• Angiographic catheterization of specific coronary artery (e.g LAD)
2. Rapid pacing models (Tachycardia induced)
3. Valvular models
• Mitral regurgitation (MR)
4. Deoxy-corticosterone acetate (DOCA)–salt rodent model for pig
5. Pressure overload models
• Aortic constriction in pigs
• Porcine model of progressive LV pressure overload (LVPO)

86. VIII. Swine Models for Heart Failure
1. Balloon occlusion of LAD coronary artery
• Closed-chest techniques are preferable to open-chest techniques because
closed-chest techniques produce a swine model with intact anatomical
structures that are more similar to those found in humans
• Coronary angioplasty balloon catheter is used
• Closed-chest balloon occlusion for 90 min followed by balloon deflation and
removal, resulting in complete reperfusion of the coronary artery—hence, an
open-artery or ischemia-reperfusion model in the closed chest (fig on next slide)
• Hemodynamics and neurohumoral activation over the first 24 h
consistent with acute MI
• Longer-term (1–4 weeks) effects to raise key hemodynamic indices of
HF

87. Balloon occlusion/reperfusion:
PTCA- Percutaneous Transluminal Coronary Angioplasty

88. VIII. Swine Models for Heart Failure
Procedure: (Balloon occlusion model)
• A right femoral arterial access- using the Seldinger technique
• A 7Fr introducer sheath place percutaneously
• Under fluoroscopic guidance- 6Fr hockey stick guiding catheter
• Introduce and place at the origin of the left coronary artery
• Coronary angiograms were obtained in the 40∘ left anterior oblique
(LAO) projection to better demonstrate the length of the left anterior
descending coronary artery (LAD) and a 0.014’’ guidewire advanced
inside this artery
• Over-the-wire coronary balloon of appropriate diameter (typically 3
mm) position either below the larger diagonal branch

89. VIII. Swine Models for Heart Failure
Procedure:
• The balloon inflated and correct occlusion assessed by contrast injection
through the guiding catheter
• Wire- removed and balloon lumen was flushed with 5mL of saline
before injecting 3mL of absolute ethanol at a rate of 1 mL/min
• Further 5mL of heparinized saline administered before balloon deflation
and removal
• A postprocedural coronary angiogram obtained
• Animals are kept under anesthesia with lidocaine infusion for another
hour, before being sent for an immediate MR follow-up study
• Explanted hearts were submerged in 4% formalin for a minimum of 48
hours

90. VIII. Swine Models for Heart Failure
2. Rapid pacing models

91. VIII. Swine Models for Heart Failure
3. Swine Model of Mitral Regurgitation Induced Heart
Failure[20]
• Inducing MR by severing the mitral valve chordae tendinae in pigs
• Using two different percutaneous approaches
• Antegrade approach method uses transseptal puncture to access the mitral
valve chords through the LA
• Retrograde approach accesses the mitral valve cords from the aorta side via
carotid arterial access

92. VIII. Swine Models for Heart Failure
• Other closed-chest techniques include:
• Intracoronary administration of ethanol
• Embolization using coils or microbeads

93. XI. Transgenic Mice

94. 6. Transgenic Mice
Several genetic models of heart failure
By addition or deletion of genes in mice
Miniaturized physiological techniques to evaluate the resulting cardiac
phenotypes
These models allow the identification of genes that are causative for
heart failure and to evaluate the molecular mechanisms responsible for
the development and progression of the disease
Developed

95. 6. Transgenic Mice
Gene targeted disruption of the muscle LIM protein (MLP) in
mice- New Model for heart failure
• MLP is a regulator of myogenic differentiation
• Mice who were homozygous for the MLP knockout develop dilated
cardiac myopathy (myocardial hypertrophy)
• Adult mice show clinical and hemodynamic signs of heart failure similar
to those in humans

96. 6. Transgenic Mice
• A recent Model of transgenic overexpression of tropomodulin
• Exhibited dilated cardiomyopathy 2–4 weeks after birth with reduced
contractile function and heart failure
• This was associated with the loss of myofibrillar organization.
Tested Group Control Group
Drug administered orally
SC or IP for 15 days
Not Treated
Compared at the end of the experiment
Not Treated
Animals

97. X. Other Models for Heart Failure

98. X. Other Animal Models for HF
1. Cryoinjury model:
• Used to study Myocardial Ischemia
• Often used for mice and zebrafish
• This model can be employed in larger mammals, such as pig
2. Hydraulic occluder or the ameroid ring constrictor:
• Induce myocardial ischemia/infarction
• Occluder is inflated in order to induce either partial stenosis or complete
occlusion
• This model mainly for large animals, such as pigs
3. Profilin-1 transgenic mice:
• Models of overload-induced heart failure
• Pharmacological goals for treatment of heart failure and prospective

99. X. Other Animal Models for HF
4. Other Mice Models for Ischaemic injury:
• 129S6 mice- Infarct rupture was most frequently observed
• Swiss mice- Cardiac dilatation was most prominent
5. HF-DB mouse model: (Heart failure- De-banding)
• Recently, a mouse model with combined MI and temporary TAC was developed
• which has enabled the elucidation of the impact of mechanical unloading
following ischaemic injury
• Female 8 week old C57BL/6 mice used
• Murine Model of Reversible Heart Failure

100. X. Other Animal Models for HF
6. Model for T1D is Akita mouse (Ins2Akitaþ+/-):
• To investigate the impact of Type 1 (T1D) and Type 2 diabetes (T2D) on the heart
• Gene exhibits a mutation in the Insulin2 encoding gene
• Hearts from Akita mice show increased inflammation and
• Diastolic dysfunction in the presence of normal systolic function
7. Ob/ob77 and db/db mice:
• Commonly used models of obesity and T2D
• Based on leptin resistance (Ob/ob) or deficiency (db/db)
8. Zucker fatty (ZF) rats: (express non-functional leptin receptors)
• Models for T2D and insulin resistance
• These Inbred strain of ZF rats with high serum glucose levels

101. X. Other Animal Models for HF
9. Senescence-accelerated prone (SAMP) mice:
• Generated by selective inbreeding of AKR/mice with inherited senescence
• Used to study various effects of aging
• SAMP mice develop age dependent diastolic dysfunction, adverse remodeling
• Also, HFpEF
10. Aorto-caval shunt Model:
• For Mice, Rats, Dogs and Pigs (rat is the predominant species)
• Surgery- led to increased end-diastolic and end-systolic volume and load
• Used to study isolated RV (Right Ventricle) volume overload
11. Pulmonary regurgitation model: (Pulmonary valve insufficiency)
• Predominantly in pigs but also in sheep and mice
• Created by sutures- wall of the pulmonary trunk around hinge points of the
pulmonary valve leaflets

102. X. Other Animal Models for HF
12. Sugen hypoxia (SuHx) in rat Model:
• 2-hit model
• Single SC injection with VEGF-receptor antagonist Sugen 5416 is followed by 3–4
weeks of hypoxia
• Causes endothelial hyperproliferation leads to progressive pulmonary vascular
occlusion- Cause Pulmonary Hypertension (PH)
13. Sugen hypxoxia in mice Model:
• Weekly Sugen 5416 injections during exposure to hypoxia cause PH in mice

103. X. Other Animal Models for HF
14. Chronic Thromboembolic Pulmonary Hypertension (CTEPH) models:
• One model of CTEPH ligate the left pulmonary artery followed by weekly
injection of histoacryl in the artery of the right lower lobe for 5 weeks
• Another model induces a percutaneously placed cobber scaffold followed by
embolization and tranexamic acid
15. Genetic models for RV failure and PH: include
• Bone Morphogenetic Peptide Receptor type 2 (BMPR-2) knockout mice
• Low-density lipoprotein Receptor-related Protein 1 (LRP1) deficient mice
• Insulin-resistant male apoE- deficient mice
• TGF-β1 transgenic mice
RV- Right Ventricle
PH- Pulmonary Hypertension

104. X. Other Animal Models for HF
16. To Study- Hypertrophic cardiomyopathy:
• Two new transgenic strains
• One by ablating cMyBP-C (e.g. cMyBP-C−/− mice)
• Another by overexpressing human myotrophin gene (i.e. Tg mice)
• Both shown the development of hypertrophy that progressively led to HF
17. Models of autoimmune cardiomyopathy (AICM):
• Model is engineered by crossing 2 different transgenic mice
• DQ8 transgenic non-obese diabetic (NOD) mouse with and NOD Major
Histocompatibility Complex (MHC) class II β-chain knockout (KO) line
• Which leads to premature death through development of progressive Dilated
cardiomyopathy (DCM) and HF
18. Double knock-out dystrophin/utrophin mouse model:
• Cause severe cardiac dysfunction at 8 weeks of age- HF

105. X. Other Animal Models for HF
19. New rat model of abdominal venous congestion: [8]
• They opted to constrict the (inferior vena cava) IVC in the thoracic cavity in an
easy-accessible rat model
• A permanent constriction above the diaphragm was applied by tying a surgical
wire around the IVC
• This rat model offers the unique possibility of studying abdominal venous
congestion in heart failure and by extension in the cardiorenal syndrome

106. X. Other Animal Models for HF
20. Induced pluripotent stem cells (iPSCs) Model: [6]
• Adding different biochemical factors to the culture medium like:
• Triiodothyronine (T3), Ascorbic acid and Neuregulin-1β
• Having essential role in cardiac development
• Tailoring surface topography is another strategy to induce cardiomyocytes
alignment
I) 2D in vitro models-
• Pharmaceutical companies have been utilizing 2D cardiac in vitro models to
assess functional properties and test cardiotoxicity in preclinical stages for
decades
• For e.g., Electrophysiology and Rhythm disorders are among the main
parameters that have been measured in 2D models

107. X. Other Animal Models for HF
20. Induced pluripotent stem cells (iPSCs) Model: [6]
II) 3D in vitro models-
• To mimic the physiological and anatomical structure of the native heart,
researchers have used various techniques to build up more complex 3D
microenvironments
• Four main approaches, used to fabricate 3D scaffolds in vitro:
1. Encapsulating cells inside hydrogels
2. Seeding cells into prefabricated structures
3. Utilizing decellularized ECM of the native heart tissue
4. Overlaying 2D cell sheets on top of each other

108. X. Other Animal Models for HF
20. Induced pluripotent stem cells (iPSCs) Model: [6]
II) 3D in vitro models-
• Cell encapsulation inside a hydrogel is currently a main approach for creating
engineered heart tissue (EHT)
• Figures on next slide
Biowire, with a more mature phenotype and a closer recapitulation
of the adult native cardia tissue, is considered to have more predictive
power for cardiotoxicity screening
Biowire developed by seeding cardiac cells encapsulated in a hydrogel into a
polydimethylsiloxane (PDMS) microwell, where the cardiac cell suspension self-
organized around a suture template situated in the middle of the microwell

109. X. Other Animal Models for HF
• Different approaches to
fabricate engineered
heart/cardiac tissue:
A. Organ on chip
B. Cardiac patch; scale bar: 2.5mm
C. Circular EHT to apply mechanical
stimulation
D. Rod-shaped Biowire to apply
electrical stimulation; scale bar:
0.5mm

110. X. Other Animal Models for HF
• Summary of current in vitro models
CM- Cardiomyocytes; hESC- human embryonic stem cells; iPSCs- Induced pluripotent
stem cells; hESCs- Human embryonic stem cells

111. X. Other Animal Models for HF
21. Heart Transplant Models:
• Porcine Model of Orthotopic Heart Transplantation- [15]
• Video link- https://www.jove.com/video/59197/ (with full procedure)
• Heterotopic Cervical Heart Transplantation Model in Mice- [16]
• Video link- http://www.jove.com/video/52907/
• Submandibular Gland-preserving Technique for Heterotopic Cervical
Heart Transplantation in Mice- [17]

112. X. Other Animal Models for HF
22. Instrument application Models:
• Swine models of stent application
• To studying biological reactions to implanted prostheses or when testing the
clinical application of medical devices
• Normocholesterolemic domestic crossbred pig is generally preferred for
relatively short term studies
• Suited for testing the application of stents in both coronary and peripheral
arteries (same for rats)

113. In-Vitro Models

114. Animal models for Heart Failure-
In-vitro models
In vitro models:
1. Isolated hamster cardiomyopathic heart
2. Isolated cat papillary muscle
3. Ouabain binding

115. 1. Isolated hamster cardiomyopathic heart
Purpose and Rationale:
• Isolated Syrian hamster hearts can be used for evaluation of cardiotonic
drugs
Procedure:
• Syrian hamsters, age 50 weeks
• Normal as control and tests with cardiomyopathy
• Pretreated: Heparin IP (5mg/kg)
• 20 min later, heart prepared according to Langendroff method
• Perfuse with Ringer’s solution
• Allow to equilibrate for 60min at 32 C with preload of 1.5g

116. 1. Isolated hamster cardiomyopathic heart
Evaluation:
• Transducer attached to polygraph- Measured Force of contraction
• Elecroflowmeter-Measured heart Rate
• Test drugs are injected through the aortic cannula into the in flowing
heart Ringer’s solution
• Contractile force and coronary flow in heart of treated and control
group are compared using student’s t test
• % improvement is calculated

117. 2. Isolated cat papillary muscle
Purpose and Rationale
• Prolonged electrical stimulation on cardiac tissue results in decrease in
performance
• Cardiac glycoside restore the force of contraction
Procedure
• Cats either of sex, 2.5 to 3kgs are anaesthetised
• Left thoracotomy done- Heart exposed
• Papillary muscles from right ventricle are isolated and fixed in Ringer’s
at 37C
• Electrical stimulus of 4-6V are applied at 30/min and contraction are
recorded
Normal
Test

118. 2. Isolated cat papillary muscle
Evaluation
• On electrical stimulation for 1 hour muscle contraction start decreasing
• Cardiac glycoside added- restore contractile force
• Ouabain 300ng/ml
• Evaluation is based on increase in contractile force on adding glycoside
• Calculated as % of predose levels and compared between groups

119. 3. Ouabain Binding
Purpose and Rationale:
• The binding kinetics of Ouabain are similar to cardiac glycoside
Purpose
• Rats heart are submitted through coronary perfusion
• Myocytes are isolated by collagen digestion- Myocyte sarcolemma
• Radioactive ouabain (3H) with specific activity of 20Ci/mmol is
incubated with ligands at 37C for 10min
• Association process- 10/100nM ouabain + 200Ug membrane
preparation initiate the reaction

120. 3. Ouabain Binding
Procedure
1. Equilibrium binding:
• Carried out in the presence of increasing conc of (3H) ouabain(10nM to
3UM)
• 40Ug of membranes are added
• After 30 mins duplicate aliquots of 4.5ml removed and filtered
2. Dissociation Process:
• Experimental conditions are used to study association
• 10ml of pre warmed Mg2+ and Pi Tris-HCL added to 0.2mM unlabeled
ouabain to initiate dissociation process

121. 3. Ouabain Binding
Evaluation
• Radioactive bound to the filters and specific binding measures are
determined
• Kinetic parameters of association and dissociation calculated
• Data analysed by Scatchard plots

122. Summary on the
basis of Stressors
Abbreviation in fig:
DOX- doxorubicin;
EtOH- ethanol;
MCT- monocrotaline;
Hcy- homocysteine;
I/R- ischaemia/reperfusion
injury;
ISO- isoproterenol;
LAD- left anterior descending
artery;
LV- left ventriclular;
RV- right ventriclular

123. Large Animal models

124. Overview
of Large
Animal
Models

125. Small animal models of heart failure
Continued

126. Small animal models of heart failure
Continued

127. Small animal models of heart failure
Continued

128. Small animal models of heart failure

129. Selection of similarities and
differences between humans and
animal models frequently used in
cardiovascular research
(Table on next slide)

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136. Thank You
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“Making scientific and authentic drug information accessible”
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