Heme oxygenase (HO)-1 converts heme to bilirubin, carbon monoxide, and iron. Our prior work has suggested a cardioprotective role for HO-1 in heart failure. To test whether HO-1 (heat shock protein 32) prevents cardiomyocyte apoptosis and cardiac dysfunction after ischemia-reperfusion (I/R), we generated transgenic mice overexpressing HO-1 in the heart under the control of the alpha-myosin heavy chain promoter. HO-1 transcript and protein increased markedly in the heart only. In an isolated heart preparation, we observed an enhanced functional recovery during reperfusion after ischemia in the transgenic hearts compared with nontransgenic controls. I/R injury was also performed in intact animals by coronary ligation and reperfusion to assess the protective role of HO-1 overexpression on heart apoptosis. HO-1 overexpression reduced cardiac apoptosis, as evidenced by fewer terminal deoxynucleodidyl transferase-mediated dUTP nick-end labeling-positive or in situ oligo ligation-positive myocytes, compared with nontransgenic mice. Our results indicate that cardioselective overexpression of HO-1 exerts a cardioprotective effect after myocardial I/R in mice, and this effect is probably mediated via an antiapoptotic action of HO-1.

Peroxisome proliferator-activated receptors (PPAR) decrease the production of cytokine and inducible nitric-oxide synthase (iNOS) expression, which are associated with aging-related inflammation and insulin resistance. Recently, the involvement of the induction of heme oxygenase-1 (HO-1) in regulating inflammation has been suggested, but the exact mechanisms for reducing inflammation by HO-1 remains unclear. We found that overexpression of HO-1 and [Ru(CO) 3 Cl 2 ] 2 , a carbon monoxide (CO)-releasing compound, increased not only ERK5 kinase activity, but also its transcriptional activity measured by luciferase assay with the transfection of the Gal4-ERK5 reporter gene. This transcriptional activity is required for coactivation of PPAR␦ by ERK5 in C2C12 cells. [Ru(CO) 3 Cl 2 ] 2 activated PPAR␦ transcriptional activity via the MEK5/ERK5 signaling pathway. The inhibition of NF-B activity by ERK5 activation was reversed by a dominant negative form of PPAR␦ suggesting that ERK5/PPAR␦ activation is required for the anti-inflammatory effects of CO and HO-1. Based on these data, we propose a new mechanism by which CO and HO-1 mediate anti-inflammatory effects via activating ERK5/PPAR␦, and ERK5 mediates CO and HO-1-induced PPAR␦ activation via its interaction with PPAR␦.Muscle wasting is a major feature of the cachexia associated with diverse pathologies such as cancer, sepsis, diabetes, and aging (1). Several cytokines have been implicated in the pathogenesis of muscle wasting, most notably TNF-␣, 2 a pro-inflammatory cytokine that was originally called "cachectin" (1). In addition, aging-related chronic low grade inflammation by TNF-␣ plays an important role in insulin resistance (2). It has been proposed that chronic inflammation by TNF-␣-mediated NF-B activation and subsequent inducible nitric-oxide synthase (iNOS) induction relates to muscle wasting and insulin resistance as we will explain below. Cai et al. (3) have shown that activation of NF-B, through muscle-specific transgenic expression of activated IB kinase ␤ (MIKK), causes profound muscle wasting that resembles clinical cachexia. In contrast, no overt phenotype was seen upon muscle-specific inhibition of NF-B through expression of IB suppressor (MISR), and denervation and tumor-induced muscle loss were substantially reduced and survival rates improved by NF-B inhibition in MISR mice, which is consistent with a critical role for NF-B in the pathology of muscle wasting, especially in diabetes and during the process of aging (3). Recent studies suggest the involvement of iNOS in the pathogenesis of insulin resistance (4, 5). First, most inducers of insulin resistance, including obesity (6), free fatty acids (7), hyperglycemia (8, 9), TNF-␣, oxidative stress, and endotoxin, increase iNOS expression. Second, iNOS mediates the impaired insulin-stimulated glucose uptake by treatment with TNF-␣ and lipopolysaccharide in cultured muscle cells (10). iNOS expression is elevated in skeletal muscle of patients with type 2 diabetes (11, 12), and high fat diet-induced...

Small intestinal crypt cells play a critical role in modulating Cl- secretion during digestion. The types of Cl- channels mediating Cl- secretion in the small intestine was investigated using the intestinal epithelial cell line, IEC-18, which was derived from rat small intestine crypt cells. In initial radioisotope efflux studies, exposure to forskolin, ionomycin or a decrease in extracellular osmolarity significantly increased 36Cl efflux as compared to control cells. Whole cell patch clamp techniques were subsequently used to examine in more detail the swelling-, Ca2+-, and cAMP-activated Cl- conductance. Decreasing the extracellular osmolarity from 290 to 200 mOsm activated a large outwardly rectifying Cl- current that was voltage-independent and had an anion selectivity of I- > Cl-. Increasing cytosolic Ca2+ by ionomycin activated whole cell Cl- currents, which were also outwardly rectifying but were voltage-dependent. The increase in intracellular Ca2+ levels with ionomycin was confirmed with fura-2 loaded IEC-18 cells. A third type of whole cell Cl- current was observed after increases in intracellular cAMP induced by forskolin. These cAMP-activated Cl- currents have properties consistent with cystic fibrosis transmembrane regulator (CFTR) Cl- channels, as the currents were blocked by glibenclamide or NPPB but insensitive to DIDS. In addition, the current-voltage relationship was linear and had an anion selectivity of Cl- > I-. Confocal immunofluorescence studies and Western blots with two different anti-CFTR antibodies confirmed the expression of CFTR. These results suggest that small intestinal crypt cells express multiple types of Cl- channels, which may all contribute to net Cl- secretion.