*2.5. Hypertrophy Induced by Stimulation with Pro-Hypertrophic Factors*

KLF15 mRNA expression is low in NRVM but induced with serum starvation with an expression pattern that is antiparallel to that of ANP and BNP gene expression [13]. Treatment of NRVM with the pro-hypertrophic stimuli, endothelin-1 and phenylephrine, decreased KLF15 mRNA expression and induced ANP and BNP expression. Similarly, in cultured neonatal rat cardiomyocytes, stimulation with phenylephrine, endothelin-1 and TGFβ significantly reduced KLF15 expression levels [15]. However KLF15 expression was unchanged when cardiomyocyte growth was induced with physiological hypertrophy stimuli (i.e., insulin, insulin-like growth factor 1 or 2). These results support a role for the involvement of KLF15 in pathological hypertrophy but not in physiological hypertrophy. TGFβ-mediated down regulation of KLF15 was abolished with knockdown of the TGFβ receptor 1 [15]. TGFβ-mediated activation of p38 mitogen-activated protein kinase was also necessary and sufficient to decrease KLF15 expression. Adenoviral overexpression of an upstream p38 kinase (MKK6) that induced increased p38 phosphorylation led to an 80% decrease in KLF15 mRNA and the induction of BNP. No other studies have explored TGFβ stimulated hypertrophy and KLF15 levels in cardiomyocytes.

#### **3. Human Studies of KLF15**

#### *3.1. KLF15 Expression in Human Cardiac Tissue*

Consistent with the observation in experimental studies, there is evidence that loss of cardiac KLF15 expression may contribute to pathological LVH in humans (Table 3). Three separate studies investigated KLF15 expression in LV tissue obtained from patients undergoing cardiac surgery. In patients with LVH secondary to aortic stenosis, KLF15 protein was reduced in myocardial needle biopsy samples taken from the anterior LV of patients undergoing open heart surgery (*n* = 8) compared to patients undergoing coronary bypass grafting (*n* = 6) [13]. These patients were selected from a larger group of patients from an earlier study [21]. The characteristics of the subgroup of patients used in the paper by Fisch et al. [13] were not provided. KLF15 mRNA was also significantly reduced in LV samples of patients with non-ischaemic cardiomyopathy (*n* = 36) compared to control tissue from non-failing hearts deemed unsuitable for transplantation (*n* = 30) [10].

In patients with end-stage heart failure undergoing implantation and explantation of a left ventricular assist device (LVAD) as a bridge to transplantation, pre-device implantation LV KLF15 mRNA expression was reduced in the failing heart compared to control non-failing hearts, with significant recovery of KLF15 expression after mechanical unloading [14]. The latter study used 3–4 samples per group, collected as part of a larger study (36 pre-LVAD, 30 post-LVAD) [22].
