Gene therapy with phosphodiesterases 2A and 4B ameliorates heart failure and arrhythmias by improving subcellular cAMP compartmentation.

AIMS: Gene therapy with cardiac phosphodiesterases (PDEs) such as PDE4B has recently been described to effectively prevent heart failure in mice. However, exact molecular mechanisms of its beneficial effects, apart from general lowering of cardiomyocyte cyclic adenosine monophosphate (cAMP) levels, have not been elucidated. Here we studied whether gene therapy with two types of PDEs, namely PDE2A and PDE4B, can prevent pressure-overload induced heart failure in mice by acting on and restoring altered cAMP compartmentalization in distinct subcellular microdomains. METHODS AND RESULTS: Heart failure was induced by transverse aortic constriction followed by tail-vein injection of adeno-associated-virus type 9 vectors to overexpress PDE2A3, PDE4B3 or luciferase for 8 weeks. Heart morphology and function was assessed by echocardiography and histology which showed that PDE2A and especially PDE4B gene therapy could attenuate cardiac hypertrophy, fibrosis and decline of contractile function. Live cell imaging using targeted cAMP biosensors showed that PDE overexpression restored altered cAMP compartmentalization in microdomains associated with ryanodine receptor type 2 (RyR2) and caveolin-rich plasma membrane. This was accompanied by ameliorated caveolin-3 decline after PDE2A3 overexpression, reduced RyR2 phosphorylation in PDE4B3 overexpressing hearts and antiarrhythmic effects of both PDEs measured under isoproterenol stimulation in single cells. Strong association of overexpressed PDE4B but not PDE2A with RyR2 microdomain could prevent calcium leak and arrhythmias in human induced pluripotent stem derived cardiomyocytes with the A2254 V mutation in RyR2 causing catecholaminergic polymorphic ventricular tachycardia. CONCLUSIONS: Our data indicate that gene therapy with phosphodiesterases can prevent heart failure including associated cardiac remodeling and arrhythmias by restoring altered cAMP compartmentalization in functionally relevant subcellular microdomains.

The role of compartmentalized ß-AR/cAMP signaling in the regulation of lipolysis in white and brown adipocytes.

White and brown adipocytes are central mediators of lipid metabolism and thermogenesis, respectively. Their function is tightly regulated by all three ß-adrenergic receptor (ß-AR) subtypes which are coupled to the production of the second messenger 3',5'-cyclic adenosine monophosphate (cAMP). While known for decades in other cell types, compartmentation of adipocyte ß-AR/cAMP signaling by spatial organization of the pathway and by cAMP degrading phosphodiesterases (PDEs) as well as its role in the regulation of lipolysis is only beginning to emerge. Here, we provide a short overview of recent findings which shed light on compartmentalized signaling using live cell imaging of cAMP in adipocytes and discuss possible future directions of research which could open up new avenues for the treatment of metabolic disorders.

Amyloid beta 42 alters cardiac metabolism and impairs cardiac function in male mice with obesity.

There are epidemiological associations between obesity and type 2 diabetes, cardiovascular disease and Alzheimer's disease. The role of amyloid beta 42 (Aß42) in these diverse chronic diseases is obscure. Here we show that adipose tissue releases Aß42, which is increased from adipose tissue of male mice with obesity and is associated with higher plasma Aß42. Increasing circulating Aß42 levels in male mice without obesity has no effect on systemic glucose homeostasis but has obesity-like effects on the heart, including reduced cardiac glucose clearance and impaired cardiac function. The closely related Aß40 isoform does not have these same effects on the heart. Administration of an Aß-neutralising antibody prevents obesity-induced cardiac dysfunction and hypertrophy. Furthermore, Aß-neutralising antibody administration in established obesity prevents further deterioration of cardiac function. Multi-contrast transcriptomic analyses reveal that Aß42 impacts pathways of mitochondrial metabolism and exposure of cardiomyocytes to Aß42 inhibits mitochondrial complex I. These data reveal a role for systemic Aß42 in the development of cardiac disease in obesity and suggest that therapeutics designed for Alzheimer's disease could be effective in combating obesity-induced heart failure.

Remodelling of cAMP dynamics within the SERCA2a microdomain in heart failure with preserved ejection fraction caused by obesity and type 2 diabetes.

AIMS: Despite massive efforts, we remain far behind in our attempts to identify effective therapies to treat heart failure with preserved ejection fraction (HFpEF). Diastolic function is critically regulated by sarcoplasmic/endoplasmic reticulum (SR) calcium ATPase 2a (SERCA2a), which forms a functional cardiomyocyte (CM) microdomain where 3',5'-cyclic adenosine monophosphate (cAMP) produced upon ß-adrenergic receptor (ß-AR) stimulation leads to phospholamban (PLN) phosphorylation and facilitated Ca2+ re-uptake. METHODS AND RESULTS: To visualize real-time cAMP dynamics in the direct vicinity of SERCA2a in healthy and diseased myocytes, we generated a novel mouse model on the leprdb background that stably expresses the Epac1-PLN Förster resonance energy transfer biosensor. Mice homozygous for the leprdb mutation (db/db) developed obesity and type 2 diabetes and presented with a HFpEF phenotype, evident by mild left ventricular hypertrophy and elevated left atria filling pressures. Live cell imaging uncovered a substantial ß2-AR subtype stimulated cAMP response within the PLN/SERCA2a microdomain of db/db but not healthy control (db/+) CMs, which was accompanied by increased PLN phosphorylation and accelerated calcium re-uptake. Importantly, db/db CMs also exhibited a desensitization of ß1-AR stimulated cAMP pools within the PLN/SERCA2a microdomain, which was accompanied by a blunted lusitropic effect, suggesting that the increased ß2-AR control is an intrinsic compensatory mechanism to maintain PLN/SERCA2a-mediated calcium dynamics and cardiac relaxation. Mechanistically, this was due to a local loss of cAMP-degrading phosphodiesterase 4 associated specifically with the PLN/SERCA2a complex. CONCLUSION: These newly identified alterations of cAMP dynamics at the subcellular level in HFpEF should provide mechanistic understanding of microdomain remodelling and pave the way towards new therapies.

CRISPR/Cas9 Knock-Out in Primary Neonatal and Adult Cardiomyocytes Reveals Distinct cAMP Dynamics Regulation by Various PDE2A and PDE3A Isoforms.

Cyclic nucleotide phosphodiesterases 2A (PDE2A) and PDE3A play an important role in the regulation of cyclic adenosine monophosphate (cAMP) and cyclic guanosine monophosphate (cGMP)-to-cAMP crosstalk. Each of these PDEs has up to three distinct isoforms. However, their specific contributions to cAMP dynamics are difficult to explore because it has been challenging to generate isoform-specific knock-out mice or cells using conventional methods. Here, we studied whether the CRISPR/Cas9 approach for precise genome editing can be used to knock out Pde2a and Pde3a genes and their distinct isoforms using adenoviral gene transfer in neonatal and adult rat cardiomyocytes. Cas9 and several specific gRNA constructs were cloned and introduced into adenoviral vectors. Primary adult and neonatal rat ventricular cardiomyocytes were transduced with different amounts of Cas9 adenovirus in combination with PDE2A or PDE3A gRNA constructs and cultured for up to 6 (adult) or 14 (neonatal) days to analyze PDE expression and live cell cAMP dynamics. A decline in mRNA expression for PDE2A (~80%) and PDE3A (~45%) was detected as soon as 3 days post transduction, with both PDEs being reduced at the protein level by >50-60% in neonatal cardiomyocytes (after 14 days) and >95% in adult cardiomyocytes (after 6 days). This correlated with the abrogated effects of selective PDE inhibitors in the live cell imaging experiments based on using cAMP biosensor measurements. Reverse transcription PCR analysis revealed that only the PDE2A2 isoform was expressed in neonatal myocytes, while adult cardiomyocytes expressed all three PDE2A isoforms (A1, A2, and A3) which contributed to the regulation of cAMP dynamics as detected by live cell imaging. In conclusion, CRISPR/Cas9 is an effective tool for the in vitro knock-out of PDEs and their specific isoforms in primary somatic cells. This novel approach suggests distinct regulation of live cell cAMP dynamics by various PDE2A and PDE3A isoforms in neonatal vs. adult cardiomyocytes.

Live-cell imaging identifies cAMP microdomains regulating ß-adrenoceptor-subtype-specific lipolytic responses in human white adipocytes.

Lipolysis of stored triglycerides is stimulated via ß-adrenergic receptor (ß-AR)/3',5'-cyclic adenosine monophosphate (cAMP) signaling and inhibited via phosphodiesterases (PDEs). In type 2 diabetes, a dysregulation in the storage/lipolysis of triglycerides leads to lipotoxicity. Here, we hypothesize that white adipocytes regulate their lipolytic responses via the formation of subcellular cAMP microdomains. To test this, we investigate real-time cAMP/PDE dynamics at the single-cell level in human white adipocytes with a highly sensitive florescent biosensor and uncover the presence of several receptor-associated cAMP microdomains where cAMP signals are compartmentalized to differentially regulate lipolysis. In insulin resistance, we also detect cAMP microdomain dysregulation mechanisms that promote lipotoxicity, but regulation can be restored by the anti-diabetic drug metformin. Therefore, we present a powerful live-cell imaging technique capable of resolving disease-driven alterations in cAMP/PDE signaling at the subcellular level and provide evidence to support the therapeutic potential of targeting these microdomains.

Understanding the Role of SERCA2a Microdomain Remodeling in Heart Failure Induced by Obesity and Type 2 Diabetes.

Obesity and type 2 diabetes (T2D) are on trend to become a huge burden across all ages. They cause harm to almost every organ, especially the heart. For decades, the incidence of heart failure with impaired diastolic function (or called heart failure with preserved ejection fraction, HFpEF) has increased sharply. More and more studies have uncovered obesity and T2D to be closely associated with HFpEF. The sarcoplasmic/endoplasmic reticulum calcium ATPase2a (SERCA2a) microdomain is a key regulator of calcium reuptake into the sarcoplasmic reticulum (SR) during diastole. 3',5'-cyclic adenosine monophosphate (cAMP) and its downstream effector cAMP dependent protein kinase (PKA) act locally within the SERCA2a microdomain to regulate the phosphorylation state of the small regulatory protein phospholamban (PLN), which forms a complex with SERCA2a. When phosphorylated, PLN promotes calcium reuptake into the SR and diastolic cardiac relaxation by disinhibiting SERCA2a pump function. In this review, we will discuss previous studies investigating the PLN/SERCA2a microdomain in obesity and T2D in order to gain a greater understanding of the underlying mechanisms behind obesity- and T2D-induced diastolic dysfunction, with the aim to identify the current state of knowledge and future work that is needed to guide further research in the field.

Polycystic ovarian syndrome increases prevalence of concentric hypertrophy in normotensive obese women.

BACKGROUND: It remains unclear as to whether polycystic ovary syndrome (PCOS) is an additional risk factor in the development of left ventricular (LV) hypertrophy in obese women. In the current study, we provide clarity on this issue by rigorously analysing patient LV geometry beyond the basic clinical measures currently used. Importantly, the cohort contained only normotensive patients that would normally be deemed low risk with no further intervention required. METHODS: The study comprised 24 obese women with PCOS and 29 obese Control women. Transthoracic echocardiography was used to evaluate LV structure/function. Basic clinical and metabolic data were collected for each participant consisting of age, BMI, blood pressure, fasting glucose, LDL-C, HLD-C, cholesterol and triglyceride levels. Exclusion criteria; BMI < 30 g/m2, type 2 diabetes, hypertension. RESULTS: Both groups exhibited concentric remodelling of the LV posterior wall at a prevalence of ~20%, this associated with grade 1 diastolic dysfunction. Estimated LV mass/height2.7 was increased patients with PCOS (45 ± 2.2 vs 37 ± 1.6) with 33% exhibiting LV mass/height2.7 above ASE guidelines, compared to 7% in Controls. Furthermore, 25% of patients with PCOS were characterised with concentric hypertrophy, an alteration in LV geometry that was not observed in the Control group. CONCLUSIONS: To our knowledge, this is the first study to assess LV geometric patterns in obese women with PCOS. The results suggest that obese women with PCOS are at greater risk of concentric hypertrophy than obese only women and provide justification for additional cardiovascular risk assessment in normotensive obese/PCOS women.

Cardiac Hypertrophy Changes Compartmentation of cAMP in Non-Raft Membrane Microdomains.

3',5'-Cyclic adenosine monophosphate (cAMP) is a ubiquitous second messenger which plays critical roles in cardiac function and disease. In adult mouse ventricular myocytes (AMVMs), several distinct functionally relevant microdomains with tightly compartmentalized cAMP signaling have been described. At least two types of microdomains reside in AMVM plasma membrane which are associated with caveolin-rich raft and non-raft sarcolemma, each with distinct cAMP dynamics and their differential regulation by receptors and cAMP degrading enzymes phosphodiesterases (PDEs). However, it is still unclear how cardiac disease such as hypertrophy leading to heart failure affects cAMP signals specifically in the non-raft membrane microdomains. To answer this question, we generated a novel transgenic mouse line expressing a highly sensitive Förster resonance energy transfer (FRET)-based biosensor E1-CAAX targeted to non-lipid raft membrane microdomains of AMVMs and subjected these mice to pressure overload induced cardiac hypertrophy. We could detect specific changes in PDE3-dependent compartmentation of ß-adrenergic receptor induced cAMP in non-raft membrane microdomains which were clearly different from those occurring in caveolin-rich sarcolemma. This indicates differential regulation and distinct responses of these membrane microdomains to cardiac remodeling.

Multifaceted remodelling of cAMP microdomains driven by different aetiologies of heart failure.

Heart failure with preserved ejection fraction (HFpEF) will soon take over as the predominant form of heart failure. This is largely driven by the continuing increased incidences of obesity and type 2 diabetes (T2D), which promote HFpEF in the absence of pressure overload stresses. With beta-blockers showing little effectiveness in treating obesity/T2D HFpEF and with no HFpEF-targeted drugs currently available, we are in urgent need of a better understanding of how obesity/T2D HFpEF develops and how we may treat this condition. An exciting emerging field aiming to do this focuses on the investigation of 3',5'-cyclic adenosine monophosphate (cAMP) microdomains in the heart. The previous work has largely focused on the investigation of cAMP microdomain remodelling in heart failure with reduced ejection fraction (HFrEF), with this work uncovering potential new targets for intervention strategies that otherwise would have been overlooked when studying changes in cAMP dynamics at the whole-cell level. In this review, we aimed to discuss current advancements in our understanding of cAMP microdomain remodelling in HFrEF vs that in obesity/T2D-associated HFpEF, with particular focus on the unresolved questions and limitations we face in being able to translate this knowledge.

Loss of protein kinase D activity demonstrates redundancy in cardiac glucose metabolism and preserves cardiac function in obesity.

OBJECTIVE: Protein kinase D (PKD) signaling has been implicated in stress-induced cardiac remodeling and function as well as metabolic processes including contraction-mediated cardiac glucose uptake. PKD has recently emerged as a nutrient-sensing kinase that is activated in high-lipid environments, such as in obesity. However, the role of PKD signaling in cardiac glucose metabolism and cardiac function in both normal and obese conditions remains unknown. METHODS: A cardiac-specific and inducible dominant negative (DN) PKD mouse model was developed. Echocardiography was used to assess cardiac function, while metabolic phenotyping was performed, including stable isotope metabolomics on cardiac tissue in mice fed either regular chow or a high-fat diet (43% calories from fat). RESULTS: Cardiac PKD activity declined by ∼90% following DN PKD induction in adult mice. The mice had diminished basal cardiac glucose clearance, suggesting impaired contraction-mediated glucose uptake, but normal cardiac function. In obesity studies, systolic function indices were reduced in control mice, but not in cardiac DN PKD mice. Using targeted stable isotope metabolomic analyses, no differences in glucose flux through glycolysis or the TCA cycle were observed between groups. CONCLUSIONS: The data show that PKD contributes to cardiac dysfunction in obesity and highlight the redundancy in cardiac glucose metabolism that maintains cardiac glucose flux in vivo. The data suggest that impairments in contraction-mediated glucose uptake are unlikely to drive cardiac dysfunction in both normal and metabolic disease states.

Increased arrhythmia susceptibility in type 2 diabetic mice related to dysregulation of ventricular sympathetic innervation.

Patients with type 2 diabetes mellitus (T2DM) have a greater risk of developing life-threatening cardiac arrhythmias. Because the underlying mechanisms and potential influence of diabetic autonomic neuropathy are not well understood, we aimed to assess the relevance of a dysregulation in cardiac autonomic tone. Ventricular arrhythmia susceptibility was increased in Langendorff-perfused hearts isolated from mice with T2DM (db/db). Membrane properties and synaptic transmission were similar at cardiac postganglionic parasympathetic neurons from diabetic and control mice; however, a greater asynchronous neurotransmitter release was present at sympathetic postganglionic neurons from the stellate ganglia of db/db mice. Western blot analysis showed a reduction of tyrosine hydroxylase (TH) from the ventricles of db/db mice, which was confirmed with confocal imaging as a heterogeneous loss of TH-immunoreactivity from the left ventricular wall but not the apex. In vivo stimulation of cardiac parasympathetic (vagus) or cardiac sympathetic (stellate ganglion) nerves induced similar changes in heart rate in control and db/db mice, and the kinetics of pacing-induced Ca2+ transients (recorded from isolated cardiomyocytes) were similar in control and db/db cells. Antagonism of cardiac muscarinic receptors did not affect the frequency or severity of arrhythmias in db/db mice, but sympathetic blockade with propranolol completely inhibited arrhythmogenicity. Collectively, these findings suggest that the increased ventricular arrhythmia susceptibility of type 2 diabetic mouse hearts is due to dysregulation of the sympathetic ventricular control.NEW & NOTEWORTHY Patients with type 2 diabetes mellitus have greater risk of suffering from sudden cardiac death. We found that the increased ventricular arrhythmia susceptibility in type 2 diabetic mouse hearts is due to cardiac sympathetic dysfunction. Sympathetic dysregulation is indicated by an increased asynchronous release at stellate ganglia, a heterogeneous loss of tyrosine hydroxylase from the ventricular wall but not apex, and inhibition of ventricular arrhythmias in db/db mice after ß-sympathetic blockade.

APP deficiency results in resistance to obesity but impairs glucose tolerance upon high fat feeding.

The amyloid precursor protein (APP) generates a number of peptides when processed through different cleavage mechanisms, including the amyloid beta peptide that is implicated in the development of Alzheimer's disease. It is well established that APP via its cleaved peptides regulates aspects of neuronal metabolism. Emerging evidence suggests that amyloidogenic processing of APP can lead to altered systemic metabolism, similar to that observed in metabolic disease states. In the present study, we investigated the effect of APP deficiency on obesity-induced alterations in systemic metabolism. Compared with WT littermates, APP-deficient mice were resistant to diet-induced obesity, which was linked to higher energy expenditure and lipid oxidation throughout the dark phase and was associated with increased spontaneous physical activity. Consistent with this lean phenotype, APP-deficient mice fed a high-fat diet (HFD) had normal insulin tolerance. However, despite normal insulin action, these mice were glucose intolerant, similar to WT mice fed a HFD. This was associated with reduced plasma insulin in the early phase of the glucose tolerance test. Analysis of the pancreas showed that APP was required to maintain normal islet and ß-cell mass under high fat feeding conditions. These studies show that, in addition to regulating aspects of neuronal metabolism, APP is an important regulator of whole body energy expenditure and glucose homeostasis under high fat feeding conditions.

Impact of maternal high fat diet on hypothalamic transcriptome in neonatal Sprague Dawley rats.

Maternal consumption of a high fat diet during early development has been shown to impact the formation of hypothalamic neurocircuitry, thereby contributing to imbalances in appetite and energy homeostasis and increasing the risk of obesity in subsequent generations. Early in postnatal life, the neuronal projections responsible for energy homeostasis develop in response to appetite-related peptides such as leptin. To date, no study characterises the genome-wide transcriptional changes that occur in response to exposure to high fat diet during this critical window. We explored the effects of maternal high fat diet consumption on hypothalamic gene expression in Sprague Dawley rat offspring at postnatal day 10. RNA-sequencing enabled discovery of differentially expressed genes between offspring of dams fed a high fat diet and offspring of control diet fed dams. Female high fat diet offspring displayed altered expression of 86 genes (adjusted P-value<0.05), including genes coding for proteins of the extra cellular matrix, particularly Collagen 1a1 (Col1a1), Col1a2, Col3a1, and the imprinted Insulin-like growth factor 2 (Igf2) gene. Male high fat diet offspring showed significant changes in collagen genes (Col1a1 and Col3a1) and significant upregulation of two genes involved in regulation of dopamine availability in the brain, tyrosine hydroxylase (Th) and dopamine reuptake transporter Slc6a3 (also known as Dat1). Transcriptional changes were accompanied by increased body weight, body fat and body length in the high fat diet offspring, as well as altered blood glucose and plasma leptin. Transcriptional changes identified in the hypothalamus of offspring of high fat diet mothers could alter neuronal projection formation during early development leading to abnormalities in the neuronal circuitry controlling appetite in later life, hence priming offspring to the development of obesity.

Complex Energy Metabolic Changes in Heart Failure With Preserved Ejection Fraction and Heart Failure With Reduced Ejection Fraction.

Alterations in cardiac energy metabolism contribute to the severity of heart failure. However, the energy metabolic changes that occur in heart failure are complex, and are dependent not only on the severity and type of heart failure present, but also on the coexistence of common comorbidities such as obesity and type 2 diabetes. In this article we review the cardiac energy metabolic changes that occur in heart failure. An emphasis is made on distinguishing the differences in cardiac energy metabolism between heart failure with preserved ejection fraction (HFpEF) and heart failure with reduced ejection fraction (HFrEF) and in clarifying the common misconceptions surrounding the fate of fatty acids and glucose in the failing heart. The major key points from this article are: (1) mitochondrial oxidative capacity is reduced in HFpEF and HFrEF; (2) fatty acid oxidation is increased in HFpEF and reduced in HFrEF (however, oxidative metabolism of fatty acids in HFrEF still exceeds that of glucose); (3) glucose oxidation is decreased in HFpEF and HFrEF; (4) there is an uncoupling between glucose uptake and oxidation in HFpEF and HFrEF, resulting in an increased rate of glycolysis; (5) ketone body oxidation is increased in HFrEF, which might further reduce fatty acid and glucose oxidation; and finally, (6) branched chain amino acid oxidation is impaired in HFrEF. The understanding of these changes in cardiac energy metabolism in heart failure are essential to allow the development of metabolic modulators in the treatment of heart failure.

Obesity and type 2 diabetes have additive effects on left ventricular remodelling in normotensive patients-a cross sectional study.

BACKGROUND: It is unclear whether obesity and type 2 diabetes (T2D), either alone or in combination, induce left ventricular hypertrophy (LVH) independent of hypertension. In the current study, we provide clarity on this issue by rigorously analysing patient left ventricular (LV) structure via clinical indices and via LV geometric patterns (more commonly used in research settings). Importantly, our sample consisted of hypertensive patients that are routinely screened for LVH via echocardiography and normotensive patients that would normally be deemed low risk with no further action required. METHODS: This cross sectional study comprised a total of 353 Caucasian patients, grouped based on diagnosis of obesity, T2D and hypertension, with normotensive obese patients further separated based on metabolic health. Basic metabolic parameters were collected and LV structure and function were assessed via transthoracic echocardiography. Multivariable logistic and linear regression analyses were used to identify predictors of LVH and diastolic dysfunction. RESULTS: Metabolically healthy normotensive obese patients exhibited relatively low risk of LVH. However, normotensive metabolically non-healthy obese, T2D and obese/T2D patients all presented with reduced normal LV geometry that coincided with increased LV concentric remodelling. Furthermore, normotensive patients presenting with both obesity and T2D had a higher incidence of concentric hypertrophy and grade 3 diastolic dysfunction than normotensive patients with either condition alone, indicating an additive effect of obesity and T2D. Alarmingly these alterations were at a comparable prevalence to that observed in hypertensive patients. Interestingly, assessment of LVPWd, a traditional index of LVH, underestimated the presence of LV concentric remodelling. The implications for which were demonstrated by concentric remodelling and concentric hypertrophy strongly associating with grade 1 and 3 diastolic dysfunction respectively, independent of sex, age and BMI. Finally, pulse pressure was identified as a strong predictor of LV remodelling within normotensive patients. CONCLUSIONS: These findings show that metabolically non-healthy obese, T2D and obese/T2D patients can develop LVH independent of hypertension. Furthermore, that LVPWd may underestimate LV remodelling in these patient groups and that pulse pressure can be used as convenient predictor of hypertrophy status.

The PKD inhibitor CID755673 enhances cardiac function in diabetic db/db mice.

The development of diabetic cardiomyopathy is a key contributor to heart failure and mortality in obesity and type 2 diabetes (T2D). Current therapeutic interventions for T2D have limited impact on the development of diabetic cardiomyopathy. Clearly, new therapies are urgently needed. A potential therapeutic target is protein kinase D (PKD), which is activated by metabolic insults and implicated in the regulation of cardiac metabolism, contractility and hypertrophy. We therefore hypothesised that PKD inhibition would enhance cardiac function in T2D mice. We first validated the obese and T2D db/db mouse as a model of early stage diabetic cardiomyopathy, which was characterised by both diastolic and systolic dysfunction, without overt alterations in left ventricular morphology. These functional characteristics were also associated with increased PKD2 phosphorylation in the fed state and a gene expression signature characteristic of PKD activation. Acute administration of the PKD inhibitor CID755673 to normal mice reduced both PKD1 and 2 phosphorylation in a time and dose-dependent manner. Chronic CID755673 administration to T2D db/db mice for two weeks reduced expression of the gene expression signature of PKD activation, enhanced indices of both diastolic and systolic left ventricular function and was associated with reduced heart weight. These alterations in cardiac function were independent of changes in glucose homeostasis, insulin action and body composition. These findings suggest that PKD inhibition could be an effective strategy to enhance heart function in obese and diabetic patients and provide an impetus for further mechanistic investigations into the role of PKD in diabetic cardiomyopathy.