Mammalian skeletal muscle is broadly characterized by the presence of two distinct categories of muscle fibers called type I "red" slow twitch and type II "white" fast twitch, which display marked differences in contraction strength, metabolic strategies, and susceptibility to fatigue. The relative representation of each fiber type can have major influences on susceptibility to obesity, diabetes, and muscular dystrophies. However, the molecular factors controlling fiber type specification remain incompletely defined. In this study, we describe the control of fiber type specification and susceptibility to metabolic disease by folliculin interacting protein-1 (Fnip1). Using Fnip1 null mice, we found that loss of Fnip1 increased the representation of type I fibers characterized by increased myoglobin, slow twitch markers [myosin heavy chain 7 (MyH7), succinate dehydrogenase, troponin I 1, troponin C1, troponin T1], capillary density, and mitochondria number. Cultured Fnip1-null muscle fibers had higher oxidative capacity, and isolated Fnip1-null skeletal muscles were more resistant to postcontraction fatigue relative to WT skeletal muscles. Biochemical analyses revealed increased activation of the metabolic sensor AMP kinase (AMPK), and increased expression of the AMPK-target and transcriptional coactivator PGC1α in Fnip1 null skeletal muscle. Genetic disruption of PGC1α rescued normal levels of type I fiber markers MyH7 and myoglobin in Fnip1-null mice. Remarkably, loss of Fnip1 profoundly mitigated muscle damage in a murine model of Duchenne muscular dystrophy. These results indicate that Fnip1 controls skeletal muscle fiber type specification and warrant further study to determine whether inhibition of Fnip1 has therapeutic potential in muscular dystrophy diseases.

Inhibition of voltage-gated, L-type Ca2+ (CaL) channels by clinical calcium channel blockers provides symptomatic improvement to some pediatric patients with pulmonary arterial hypertension (PAH). The present study investigated whether abnormalities of vascular CaL channels contribute to the pathogenesis of neonatal PAH using a newborn piglet model of hypoxia-induced PAH. Neonatal piglets exposed to chronic hypoxia (CH) developed PAH by 21 days, which was evident as a 2.1-fold increase in pulmonary vascular resistance in vivo compared with piglets raised in normoxia (N). Transpulmonary pressures (ΔPtp) in the corresponding isolated perfused lungs were 20.5 ± 2.1 mmHg (CH) and 11.6 ± 0.8 mmHg (N). Nifedipine reduced the elevated ΔPtp in isolated lungs of CH piglets by 6.4 ± 1.3 mmHg but only reduced ΔPtp in lungs of N piglets by 1.9 ± 0.2 mmHg. Small pulmonary arteries from CH piglets also demonstrated accentuated Ca2+-dependent contraction, and Ca2+ channel current was 3.94-fold higher in the resident vascular muscle cells. Finally, although the level of mRNA encoding the pore-forming α1C-subunit of the CaL channel was similar between small pulmonary arteries from N and CH piglets, a profound and persistent upregulation of the vascular α1C protein was detected by 10 days in CH piglets at a time when pulmonary vascular resistance was only mildly elevated. Thus chronic hypoxia in the neonate is associated with the anomalous upregulation of CaL channels in small pulmonary arteries in vivo and the resulting abnormal Ca2+-dependent resistance may contribute to the pathogenesis of PAH.

Hematopoietic protein-1 (Hem-1) is a hematopoietic cell specific member of the WAVE (Wiskott-Aldrich syndrome verprolin-homologous protein) complex, which regulates filamentous actin (F-actin) polymerization in many cell types including immune cells. However, the roles of Hem-1 and the WAVE complex in erythrocyte biology are not known. In this study, we utilized mice lacking Hem-1 expression due to a non-coding point mutation in the Hem1 gene to show that absence of Hem-1 results in microcytic, hypochromic anemia characterized by abnormally shaped erythrocytes with aberrant F-actin foci and decreased lifespan. We find that Hem-1 and members of the associated WAVE complex are normally expressed in wildtype erythrocyte progenitors and mature erythrocytes. Using mass spectrometry and global proteomics, Coomassie staining, and immunoblotting, we find that the absence of Hem-1 results in decreased representation of essential erythrocyte membrane skeletal proteins including α- and β- spectrin, dematin, p55, adducin, ankyrin, tropomodulin 1, band 3, and band 4.1. Hem1−/− erythrocytes exhibit increased protein kinase C-dependent phosphorylation of adducin at Ser724, which targets adducin family members for dissociation from spectrin and actin, and subsequent proteolysis. Increased adducin Ser724 phosphorylation in Hem1−/− erythrocytes correlates with decreased protein expression of the regulatory subunit of protein phosphatase 2A (PP2A), which is required for PP2A-dependent dephosphorylation of PKC targets. These results reveal a novel, critical role for Hem-1 in the homeostasis of structural proteins required for formation and stability of the actin membrane skeleton in erythrocytes.

The development of chronic hypoxia (CH)‐induced neonatal pulmonary hypertension (PH) is associated with increased production of thromboxane (TxA2) and an upregulation of L‐type calcium (CaL) channels. Thus, we hypothesized that TxA2 synthase inhibition may blunt the development of PH by mitigating the upregulation of CaL channels. Newborn piglets were exposed to 21 days of normoxia (N), CH or CH plus the TxA2 synthase inhibitor, furegrelate (oral 3 mg/kg, 3x daily). In vivo pulmonary vascular resistance index (PVRI) was 3.15 ‐fold higher in CH (104±1 WU) compared to N (33±1 WU) piglets. Furegrelate partially blunted the elevated PVRI in CH piglets (64±0.5 WU). Furegrelate also reversed the elevated transpulmonary pressure in isolated lungs of CH piglets by 66%, and blunted the overexpression of CaL channels and anomalous Ca2+‐dependent tone. Pulmonary arterial distensibility was decreased in lungs of CH compared to N piglets, indicative of vascular remodeling. Furegrelate partially restored distensibility to normal levels. Our findings suggest that pharmacological inhibition of TxA2 synthase blunts the development of hypoxia‐induced neonatal PH by preserving vascular function and structural integrity. Funded by NIH R01 HL83013

The development of chronic hypoxia (CH)‐induced neonatal pulmonary hypertension (PH) is associated with increased production of thromboxane (TxA2) and an upregulation of L‐type calcium (CaL) channels. Thus, we hypothesized that TxA2 synthase inhibition may blunt the development of PH by mitigating the upregulation of CaL channels. Newborn piglets were exposed to 21 days of normoxia (N), CH or CH plus the TxA2 synthase inhibitor, furegrelate (oral 3 mg/kg, 3x daily). In vivo pulmonary vascular resistance index (PVRI) was 3.15 ‐fold higher in CH (104±1 WU) compared to N (33±1 WU) piglets. Furegrelate partially blunted the elevated PVRI in CH piglets (64±0.5 WU). Furegrelate also reversed the elevated transpulmonary pressure in isolated lungs of CH piglets by 66%, and blunted the overexpression of CaL channels and anomalous Ca2+‐dependent tone. Pulmonary arterial distensibility was decreased in lungs of CH compared to N piglets, indicative of vascular remodeling. Furegrelate partially restored distensibility to normal levels. Our findings suggest that pharmacological inhibition of TxA2 synthase blunts the development of hypoxia‐induced neonatal PH by preserving vascular function and structural integrity. Funded by NIH R01 HL83013