Eric R. Blough

Environmental Mercury and Its Toxic Effects

Mercury exists naturally and as a man-made contaminant. The release of processed mercury can lead to a progressive increase in the amount of atmospheric mercury, which enters the atmospheric-soil-water distribution cycles where it can remain in circulation for years. Mercury poisoning is the result of exposure to mercury or mercury compounds resulting in various toxic effects depend on its chemical form and route of exposure. The major route of human exposure to methylmercury (MeHg) is largely through eating contaminated fish, seafood, and wildlife which have been exposed to mercury through ingestion of contaminated lower organisms. MeHg toxicity is associated with nervous system damage in adults and impaired neurological development in infants and children. Ingested mercury may undergo bioaccumulation leading to progressive increases in body burdens. This review addresses the systemic pathophysiology of individual organ systems associated with mercury poisoning. Mercury has profound cellular, cardiovascular, hematological, pulmonary, renal, immunological, neurological, endocrine, reproductive, and embryonic toxicological effects.

Intratracheal instillation of cerium oxide nanoparticles induces hepatic toxicity in male Sprague-Dawley rats

Background Cerium oxide (CeO2) nanoparticles have been posited to have both beneficial and toxic effects on biological systems. Herein, we examine if a single intratracheal instillation of CeO2 nanoparticles is associated with systemic toxicity in male Sprague-Dawley rats. Methods and results Compared with control animals, CeO2 nanoparticle exposure was associated with increased liver ceria levels, elevations in serum alanine transaminase levels, reduced albumin levels, a diminished sodium-potassium ratio, and decreased serum triglyceride levels (P < 0.05). Consistent with these data, rats exposed to CeO2 nanoparticles also exhibited reductions in liver weight (P < 0.05) and dose-dependent hydropic degeneration, hepatocyte enlargement, sinusoidal dilatation, and accumulation of granular material. No histopathological alterations were observed in the kidney, spleen, and heart. Analysis of serum biomarkers suggested an elevation of acute phase reactants and markers of hepatocyte injury in the rats exposed to CeO2 nanoparticles. Conclusion Taken together, these data suggest that intratracheal instillation of CeO2 nanoparticles can result in liver damage.

Aging-Associated Dysfunction of Akt/Protein Kinase B: S-Nitrosylation and Acetaminophen Intervention

Background Aged skeletal muscle is characterized by an increased incidence of metabolic and functional disorders, which if allowed to proceed unchecked can lead to increased morbidity and mortality. The mechanism(s) underlying the development of these disorders in aging skeletal muscle are not well understood. Protein kinase B (Akt/PKB) is an important regulator of cellular metabolism and survival, but it is unclear if aged muscle exhibits alterations in Akt function. Here we report a novel dysfunction of Akt in aging muscle, which may relate to S-nitrosylation and can be prevented by acetaminophen intervention. Principal Findings Compared to 6- and 27-month rats, the phosphorylation of Akt (Ser473 and Thr308) was higher in soleus muscles of very aged rats (33-months). Paradoxically, these increases in Akt phosphorylation were associated with diminished mammalian target of rapamycin (mTOR) phosphorylation, along with decreased levels of insulin receptor beta (IR-β), phosphoinositide 3-kinase (PI3K), phosphatase and tensin homolog deleted on chromosome 10 (PTEN) and phosphorylation of phosphoinositide-dependent kinase-1 (PDK1) (Ser241). In vitro Akt kinase measurements and ex vivo muscle incubation experiments demonstrated age-related impairments of Akt kinase activity, which were associated with increases in Akt S-nitrosylation and inducible nitric oxide synthase (iNOS). Impairments in Akt function occurred parallel to increases in myocyte apoptosis and decreases in myocyte size and the expression of myosin and actin. These age-related disorders were attenuated by treating aged (27-month) animals with acetaminophen (30 mg/kg body weight/day) for 6-months. Conclusions These data demonstrate that Akt dysfunction and increased S-nitrosylation of Akt may contribute to age-associated disorders in skeletal muscle and that acetaminophen may be efficacious for the treatment of age-related muscle dysfunction.

The Fischer 344/NNiaHSd X Brown Norway/BiNia is a better model of sarcopenia than the Fischer 344/NNiaHSd: a comparative analysis of muscle mass and contractile properties in aging male rat models.

Sarcopenia, characterized by profound muscle atrophy and the loss of contractile function, contributes significantly to the development of frailty and functional impairment in older age. Although present in aging humans, rat models have failed to clearly demonstrate a similar degree of this age-associated loss of muscle mass and function. This investigation compared two models of rats raised specifically for aging studies, the Fischer 344/NNiaHSd (F344/N) and the Fischer 344/NNiaHSd X Brown Norway/BiNia (F344/NXBN), and sought to determine which model provides the most accurate representation of human sarcopenia. We found that aging had no effect on F344/N muscle mass or contractile function in the extensor digitorum longus (EDL) and soleus (SOL). Conversely, in the F344/NXBN model, aging was found to decrease EDL and SOL mass and contractile function. These changes were sufficient to satisfy the proposed criteria for the diagnosis of human sarcopenia based upon muscle mass and contractile function. Results indicate that the F344/NXBN provides a better model of the alterations seen in aging human muscle than the F344/N rat model.

Sarcopenia-related apoptosis is regulated differently in fast- and slow-twitch muscles of the aging F344/N × BN rat model

Age-related decreases in muscle mass have been associated with the loss of myonuclei, possibly through a mechanism involving mitochondria. It is unclear if age-related apoptotic mechanisms vary by fiber type. Here we investigate indices of apoptosis along with the regulation of apoptotic mediators in the extensor digitorum longus (EDL) and soleus of adult (6 month), old (30 month), and very old (36 month) Fischer 344/NNiaHSD x Brown Norway/BiNia (F344/N x BN) rats. Compared to 6-month muscles, aged muscles exhibited decreases in muscle mass along with increases in the number of nuclei staining positively for DNA fragmentation. The expression of Bax, Bcl-2, caspase-3 and caspase-9 was regulated differently with aging between muscle types and in a manner not consistent with mitochondria-mediated apoptosis. To investigate the potential of calpain involvement in age-related myonuclear loss, the calpain-dependent cleavage of alpha-fodrin was examined. The proteolytic cleavage of alpha-fodrin by calpains was increased in both muscles with only the 36-month soleus exhibiting increased caspase-dependent alpha-fodrin cleavage. Taken together, these data suggest that apoptotic regulatory events differ between fiber types in the aging F344/N x BN and that mitochondrial-dependent apoptosis pathways may not play a primary role in the loss of muscle nuclei with aging.

Acetaminophen prevents aging-associated hyperglycemia in aged rats: effect of aging-associated hyperactivation of p38-MAPK and ERK1/2.

Aging‐related hyperglycemia is associated with increased oxidative stress and diminished muscle glucose transporter‐4 (Glut4) that may be regulated, at least in part, by the mitogen‐activated protein kinases (MAPK).

Impaired overload-induced hypertrophy in obese Zucker rat slow-twitch skeletal muscle

The effect of insulin resistance (IR) on the adaptation of skeletal muscle loading is not well understood. Here we examine whether the soleus muscles of the lean Zucker (LZ) and insulin-resistant obese Zucker (OZ) rat exhibit differences in their ability to undergo muscle hypertrophy following 8 wk of mechanical overload. Four-week-old male LZ (n = 5) and OZ (n = 5) rats underwent unilateral surgical ablation of the gastrocnemius muscle while the contralateral hindlimb was used as an internal control. Mechanical overload increased soleus muscle wet weight (LZ 57% and OZ 33%, respectively; P < 0.05) and average type 1 fiber cross-sectional area (LZ 32% and OZ 5%, respectively; P < 0.05) in LZ and OZ rats, while the magnitude of these increases was greater in the LZ animals (P < 0.05). The reduced degree of muscle hypertrophy observed in the OZ animals was associated with decreases in the ability of the OZ soleus muscle to phosphorylate p70s6k(Thr 389) and mTOR, while phosphorylation of p70s6k(Thr 389) was increased in the LZ overloaded soleus by 83% (P < 0.05). The amount of Tuberin/TSC2 phosphorylation, an inhibitor of mTOR, was unchanged in the LZ soleus after overload while it was increased (68.3%, P < 0.05) in OZ animals. Conversely, AMPK phosphorylation was decreased in the LZ (-22.77%, P < 0.05) but increased (57%, P < 0.05) in the OZ soleus with overload. Taken together, these data suggest that IR or other related comorbidities may impair the ability of the soleus to activate mTOR signaling and undergo load-induced muscle hypertrophy.

Submicron bioactive glass tubes for bone tissue engineering

Herein we describe a method to fabricate submicron bioactive glass tubes using sol-gel and coaxial electrospinning techniques for applications in bone tissue engineering. Heavy mineral oil and gel solution were delivered by two independent syringe pumps during the coaxial electrospinning process. Subsequently, submicron bioactive glass tubes were obtained by removal of poly(vinyl pyrrolidone) and heavy mineral oil via calcination at 600 °C for 5 h. Tubular structure was confirmed by scanning electron microscopy and transmission electron microscopy imaging. We examined the bioactivity of submicron bioactive glass tubes and fibers and evaluated their biocompatibility, using electrospun poly(ε-caprolactone) fibers--a bioinactive material--for comparison. The bioactivity of the glass tubes was examined in a simulated body fluid and they demonstrated the formation of hydroxyapatite-like minerals on both the outer and inner surfaces. In contrast, mineralization only occurred on their surface for bioactive glass solid fibers. Energy-dispersive X-ray data suggested that the bioactive glass tubes had a faster induction of mineral formation than the solid fibers. We demonstrate that the proliferation rate of mouse preosteoblastic MC3T3-E1 cells on bioactive glass tubes was comparable to that on solid fibers. We also show that bioactive glass tubes can be loaded with a model protein drug, bovine serum albumin, and that these structures exhibit delayed release properties. The bioactivity of released lysozyme can be as high as 90.9%. Taken together, these data suggest that submicron bioactive glass tubes could hold great potential for use in bone tissue engineering as well as topical drug or gene delivery.

Iron-Induced Cardiac Damage: Role of Apoptosis and Deferasirox Intervention

Excess cardiac iron levels are associated with cardiac damage and can result in increased morbidity and mortality. Here, we hypothesize that elevations in tissue iron can activate caspase-dependent signaling, which leads to increased cardiac apoptosis and fibrosis, and that these alterations can be attenuated by iron chelation. Using an iron-overloaded gerbil model, we show that increased cardiac iron is associated with reduced activation of Akt (Ser473 and Thr308), diminished phosphorylation of the proapoptotic regulator Bad (Ser136), and an increased Bax/Bcl-2 ratio. These iron-overload-induced alterations in Akt/Bad phosphorylation and Bax/Bcl-2 ratio were coupled with increased activation of the downstream caspase-9 (40/38- and 17-kDa fragments) and apoptosis executioner caspase-3 (19- and 17-kDa fragments), which were accompanied by evidence of elevated cytoskeletal α-fodrin cleavage (150- and 120-kDa fragments), discontinuity of myocardial membrane dystrophin immunoreactivity, increases in the number of terminal deoxynucleotidyl transferase dUTP nick-end labeling (TUNEL)-positive cells (nucleic DNA fragmentation), and cardiac fibrosis. We demonstrate that the administration of deferasirox, a tridentate iron chelator, is associated with diminished tissue iron deposition, attenuated activation of caspases, reduced α-fodrin cleavage, improved membrane integrity, decreased TUNEL reactivity, and attenuated cardiac fibrosis. These results suggest that the activation of caspase-dependent signaling may play a role in the development of iron-induced cardiac apoptosis and fibrosis, and deferasirox, via a reduction in cardiac tissue iron levels, may be useful for decreasing the extent of iron-induced cardiac damage.

Effects of aging and gender on muscle mass and regulation of Akt-mTOR-p70s6k related signaling in the F344BN rat model

Sarcopenia is the loss of muscle mass and strength which occurs with aging. Whether the molecular basis of sarcopenia differs with muscle type and across sex is not well understood. Here we examine how aging affects the regulation of protein kinase B (Akt), the mammalian target of rapamycin (mTOR), AMP activated kinase (AMPK), p70 ribosomal S6 kinase (p70s6k), S6 ribosomal protein (rps6) and calcineurin (CaN) in the slow soleus and fast extensor digitorum longus (EDL) muscles of 6- (adult), 30- (aged), and 36-month (very aged) male and 6- (adult), 26- (aged), and 30-month (very aged) female Fischer 344xBrown Norway (F344BN) rats. In male animals, soleus and EDL muscle to body weight ratios decreased steadily with age while in the females, losses remained unchanged after 26 months. These age-related changes in the degree of muscle atrophy across sex were associated with differences in the regulation of Akt, mTOR, and p70s6k in the slow-twitch soleus and the regulation of AMPK, 4EBP1, p70s6k and rpS6 in the fast-twitch EDL. Irrespective of muscle type, aging in both the genders was associated with increased calcineurin expression. Taken together, these data suggest that indices of protein synthesis and muscle adaptation are regulated differently with aging in different muscle types and sex.