Publications by authors named "Michaele B Manigrasso"

18 Publications

  • Page 1 of 1

An Eclectic Cast of Cellular Actors Orchestrates Innate Immune Responses in the Mechanisms Driving Obesity and Metabolic Perturbation.

Circ Res 2020 05 21;126(11):1565-1589. Epub 2020 May 21.

From the Diabetes Research Program, Division of Endocrinology, Diabetes and Metabolism, Department of Medicine (L.A., H.H.R., R.A.W., M.B.M., P.F.G., R.R., A.M.S.), NYU Grossman School of Medicine, New York.

The escalating problem of obesity and its multiple metabolic and cardiovascular complications threatens the health and longevity of humans throughout the world. The cause of obesity and one of its chief complications, insulin resistance, involves the participation of multiple distinct organs and cell types. From the brain to the periphery, cell-intrinsic and intercellular networks converge to stimulate and propagate increases in body mass and adiposity, as well as disturbances of insulin sensitivity. This review focuses on the roles of the cadre of innate immune cells, both those that are resident in metabolic organs and those that are recruited into these organs in response to cues elicited by stressors such as overnutrition and reduced physical activity. Beyond the typical cast of innate immune characters invoked in the mechanisms of metabolic perturbation in these settings, such as neutrophils and monocytes/macrophages, these actors are joined by bone marrow-derived cells, such as eosinophils and mast cells and the intriguing innate lymphoid cells, which are present in the circulation and in metabolic organ depots. Upon high-fat feeding or reduced physical activity, phenotypic modulation of the cast of plastic innate immune cells ensues, leading to the production of mediators that affect inflammation, lipid handling, and metabolic signaling. Furthermore, their consequent interactions with adaptive immune cells, including myriad T-cell and B-cell subsets, compound these complexities. Notably, many of these innate immune cell-elicited signals in overnutrition may be modulated by weight loss, such as that induced by bariatric surgery. Recently, exciting insights into the biology and pathobiology of these cell type-specific niches are being uncovered by state-of-the-art techniques such as single-cell RNA-sequencing. This review considers the evolution of this field of research on innate immunity in obesity and metabolic perturbation, as well as future directions.
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http://dx.doi.org/10.1161/CIRCRESAHA.120.315900DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7250004PMC
May 2020

Deletion of the formin Diaph1 protects from structural and functional abnormalities in the murine diabetic kidney.

Am J Physiol Renal Physiol 2018 12 22;315(6):F1601-F1612. Epub 2018 Aug 22.

Diabetes Research Program, Division of Endocrinology, Diabetes, and Metabolism, Department of Medicine, New York University School of Medicine , New York, New York.

Diaphanous 1 (DIAPH1), a member of the formin family, binds to the cytoplasmic domain of the receptor for advanced glycation end products (RAGE) and is required for RAGE signal transduction. Experiments employing genetic overexpression or deletion of Ager (the gene encoding RAGE) or its pharmacological antagonism implicate RAGE in the pathogenesis of diabetes-associated nephropathy. We hypothesized that DIAPH1 contributes to pathological and functional derangements in the kidneys of diabetic mice. We show that DIAPH1 is expressed in the human and murine diabetic kidney, at least in part in the tubulointerstitium and glomerular epithelial cells or podocytes. To test the premise that DIAPH1 is linked to diabetes-associated derangements in the kidney, we rendered male mice globally devoid of Diaph1 ( Diaph1) or wild-type controls (C57BL/6 background) diabetic with streptozotocin. Control mice received equal volumes of citrate buffer. After 6 mo of hyperglycemia, diabetic Diaph1 mice displayed significantly reduced mesangial sclerosis, podocyte effacement, glomerular basement thickening, and urinary albumin-to-creatinine ratio compared with diabetic mice expressing Diaph1. Analysis of whole kidney cortex revealed that deletion of Diaph1 in diabetic mice significantly reduced expression of genes linked to fibrosis and inflammation. In glomerular isolates, expression of two genes linked to podocyte stress, growth arrest-specific 1 ( Gas1) and cluster of differentiation 36 ( Cd36), was significantly attenuated in diabetic Diaph1 mice compared with controls, in parallel with significantly higher levels of nestin (Nes) mRNA, a podocyte marker. Collectively, these data implicate DIAPH1 in the pathogenesis of diabetes-associated nephropathy and suggest that the RAGE-DIAPH1 axis is a logical target for therapeutic intervention in this disorder.
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http://dx.doi.org/10.1152/ajprenal.00075.2018DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6336994PMC
December 2018

Small Molecule Inhibition of Ligand-Stimulated RAGE-DIAPH1 Signal Transduction.

Sci Rep 2016 Mar 3;6:22450. Epub 2016 Mar 3.

Diabetes Research Program, Department of Medicine, New York University Langone Medical Center, 550 First Avenue, New York, 10016 New York, USA.

The receptor for advanced glycation endproducts (RAGE) binds diverse ligands linked to chronic inflammation and disease. NMR spectroscopy and x-ray crystallization studies of the extracellular domains of RAGE indicate that RAGE ligands bind by distinct charge- and hydrophobicity-dependent mechanisms. The cytoplasmic tail (ct) of RAGE is essential for RAGE ligand-mediated signal transduction and consequent modulation of gene expression and cellular properties. RAGE signaling requires interaction of ctRAGE with the intracellular effector, mammalian diaphanous 1 or DIAPH1. We screened a library of 58,000 small molecules and identified 13 small molecule competitive inhibitors of ctRAGE interaction with DIAPH1. These compounds, which exhibit in vitro and in vivo inhibition of RAGE-dependent molecular processes, present attractive molecular scaffolds for the development of therapeutics against RAGE-mediated diseases, such as those linked to diabetic complications, Alzheimer's disease, and chronic inflammation, and provide support for the feasibility of inhibition of protein-protein interaction (PPI).
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http://dx.doi.org/10.1038/srep22450DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4776135PMC
March 2016

Fracture healing and lipid mediators.

Bonekey Rep 2014 2;3:517. Epub 2014 Apr 2.

Department of Biochemistry & Molecular Biology, New Jersey Medical School and Graduate School of Biomedical Sciences, Rutgers, the State University of New Jersey , Newark, NJ, USA.

Lipid mediators regulate bone regeneration during fracture healing. Prostaglandins and leukotrienes are well-known lipid mediators that regulate inflammation and are synthesized from the Ω-6 fatty acid, arachidonic acid. Cyclooxygenase (COX-1 or COX-2) and 5-lipoxygenase (5-LO) catalyze the initial enzymatic steps in the synthesis of prostaglandins and leukotrienes, respectively. Inhibition or genetic ablation of COX-2 activity impairs fracture healing in animal models. Genetic ablation of COX-1 does not affect the fracture callus strength in mice, suggesting that COX-2 activity is primarily responsible for regulating fracture healing. Inhibition of cyclooxygenase activity with nonsteroidal anti-inflammatory drugs (NSAIDs) is performed clinically to reduce heterotopic ossification, although clinical evidence that NSAID treatment impairs fracture healing remains controversial. In contrast, inhibition or genetic ablation of 5-LO activity accelerates fracture healing in animal models. Even though prostaglandins and leukotrienes regulate inflammation, loss of COX-2 or 5-LO activity appears to primarily affect chondrogenesis during fracture healing. Prostaglandin or prostaglandin analog treatment, prostaglandin-specific synthase inhibition and prostaglandin or leukotriene receptor antagonism also affect callus chondrogenesis. Unlike the Ω-6-derived lipid mediators, lipid mediators derived from Ω-3 fatty acids, such as resolvin E1 (RvE1), have anti-inflammatory activity. In vivo, RvE1 can inhibit osteoclastogenesis and limit bone resorption. Although Ω-6 and Ω-3 lipid mediators have clear-cut effects on inflammation, the role of these lipid mediators in bone regeneration is more complex, with apparent effects on callus chondrogenesis and bone remodeling.
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http://dx.doi.org/10.1038/bonekey.2014.12DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4007165PMC
June 2014

Unlocking the biology of RAGE in diabetic microvascular complications.

Trends Endocrinol Metab 2014 Jan 3;25(1):15-22. Epub 2013 Sep 3.

Diabetes Research Program, Division of Endocrinology, New York University School of Medicine, 550 First Avenue, New York, NY 10016, USA. Electronic address:

The discovery of the receptor for advanced glycation end-products (RAGE) set the stage for the elucidation of important mechanisms underpinning diabetic complications. RAGE transduces the signals of advanced glycation end-products (AGEs), proinflammatory S100/calgranulins, and high mobility group box 1 (HMGB1), and is a one of a family of receptors for lysophosphatidic acid (LPA). These ligand tales weave a theme of vascular perturbation and inflammation linked to the pathogenesis of the chronic complications of diabetes. Once deemed implausible, this concept of inflammatory cues participating in diabetic complications is now supported by a plethora of experimental evidence in the macro- and microvasculature. We review the biology of ligand-RAGE signal transduction and its roles in diabetic microvascular complications, from animal models to human subjects.
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http://dx.doi.org/10.1016/j.tem.2013.08.002DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3877224PMC
January 2014

Diabetes and vascular disease: sex matters.

Vascul Pharmacol 2012 Sep-Oct;57(2-4):78-80. Epub 2012 Jul 8.

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http://dx.doi.org/10.1016/j.vph.2012.07.001DOI Listing
January 2013

Sex differences in hypertension: contribution of the renin-angiotensin system.

Gend Med 2012 Aug 12;9(4):287-91. Epub 2012 Jul 12.

Department of Physiology and Biophysics, University of Mississippi Medical Center, 2500 N State St., Jackson, MS 39216-4505, USA.

Numerous studies have shown that female human beings exhibit lower blood pressure levels over much of their life span compared with their age-matched counterparts. This sexual dimorphism is apparent in human beings as well as most, if not all, mammals. However, after the onset of menopause blood pressure levels in women increase and become similar to those in men, suggesting an important role of sex hormones in the regulation of blood pressure. The lower blood pressure levels in premenopausal women are associated with a lower risk of development and progression of cardiovascular disease and hypertension compared with age-matched men. This clear female advantage with respect to lower incidence of cardiovascular disease no longer exists after menopause, again highlighting the importance of sex hormones in the pathophysiology of cardiovascular disease in both men and women. In fact, both estrogens and androgens have been implicated in the development of cardiovascular disease and hypertension, with estrogens, in general, being protective and androgens being detrimental. Although the exact mechanisms by which sex hormones contribute to the regulation of cardiovascular function and blood pressure are still being investigated, there is increasing evidence that modulating the activity of locally active hormonal systems is one of the major mechanisms of sex hormone actions in target organs, including the vasculature and kidneys. Indeed, several studies have demonstrated the importance of the interaction between sex hormones and the renin-angiotensin system in regulating cardiovascular function and blood pressure. Furthermore, the differential effects of estrogens and androgens on the expression and activity of the components of the renin-angiotensin system could possibly explain the sex differences in blood pressure levels and the development and progression of cardiovascular disease and hypertension.
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http://dx.doi.org/10.1016/j.genm.2012.06.005DOI Listing
August 2012

Combined inhibition of aromatase activity and dihydrotestosterone supplementation attenuates renal injury in male streptozotocin (STZ)-induced diabetic rats.

Am J Physiol Renal Physiol 2012 May 1;302(9):F1203-9. Epub 2012 Feb 1.

Department of Physiology and Biophysics, University of Mississippi Medical Center, 2500 North State St., Jackson, MS 39216, USA.

Our previous studies showed that streptozotocin (STZ)-induced diabetic male rats have increased estradiol and decreased testosterone levels that correlate with renal injury (Xu Q, Wells CC, Garman GH, Asico L, Escano CS, Maric C. Hypertension 51: 1218-1224, 2008). We further showed that either supplementing dihydrotestosterone (DHT) or inhibiting estradiol biosynthesis in these diabetic rats was only partially renoprotective (Manigrasso MB, Sawyer RT, Marbury DC, Flynn ER, Maric C. Am J Physiol Renal Physiol 301: F634-F640, 2011; Xu Q, Prabhu A, Xu S, Manigrassso MB, Maric C. Am J Physiol 297: F307-F315, 2009). The aim of this study was to test the hypothesis that the combined therapy of DHT supplementation and inhibition of estradiol synthesis would afford better renoprotection than either treatment alone. The study was performed in 12-wk-old male nondiabetic (ND), STZ-induced diabetic (D), and STZ-induced diabetic rats that received the combined therapy of 0.75 mg/day of DHT along with 0.15 mg · kg(-1) · day(-1) of an aromatase inhibitor, anastrozole (Dta), for 12 wk. Treatment with the combined therapy resulted in attenuation of albuminuria by 84%, glomerulosclerosis by 55%, and tubulointerstitial fibrosis by 62%. In addition, the combined treatment decreased the density of renal cortical CD68-positive cells by 70% and decreased protein expression of transforming growth factor-β protein expression by 60%, collagen type IV by 65%, TNF-α by 55%, and IL-6 by 60%. We conclude that the combined treatment of DHT and blocking aromatase activity in diabetic male STZ-induced diabetic rats provides superior treatment than either treatment alone in the prevention of diabetic renal disease.
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http://dx.doi.org/10.1152/ajprenal.00569.2011DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3362176PMC
May 2012

Chronic hyponatremia exacerbates multiple manifestations of senescence in male rats.

Age (Dordr) 2013 Apr 5;35(2):271-88. Epub 2012 Jan 5.

Division of Endocrinology and Metabolism, Georgetown University, Washington, DC 20007, USA.

The syndrome of inappropriate antidiuretic hormone secretion (SIADH) is frequently responsible for chronic hyponatremia in the elderly due to age-related disruption of the inhibitory component of brain osmoregulatory mechanisms. Recent research has indicated that chronic hyponatremia is associated with gait disturbances, increased falls, and bone fragility in humans, and we have found that chronic hyponatremia causes increased bone resorption and reduced bone mineral density in young rats. In this study, we used a model of SIADH to study multi-organ consequences of chronic hyponatremia in aged rats. Sustained hyponatremia for 18 weeks caused progressive reduction of bone mineral density by DXA and decreased bone ash calcium, phosphate and sodium contents at the tibia and lumbar vertebrae. Administration of 10-fold higher vitamin D during the last 8 weeks of the study compensated for the reduction in bone formation and halted bone loss. Hyponatremic rats developed hypogonadism, as indicated by slightly lower serum testosterone and higher serum FSH and LH concentrations, markedly decreased testicular weight, and abnormal testicular histology. Aged hyponatremic rats also manifested decreased body fat, skeletal muscle sarcopenia by densitometry, and cardiomyopathy manifested as increased heart weight and perivascular and interstitial fibrosis by histology. These findings are consistent with recent results in cultured osteoclastic cells, indicating that low extracellular sodium concentrations increased oxidative stress, thereby potentially exacerbating multiple manifestations of senescence. Future prospective studies in patients with SIADH may indicate whether these multi-organ age-related comorbidities may potentially contribute to the observed increased incidence of fractures and mortality in this population.
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http://dx.doi.org/10.1007/s11357-011-9347-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3592950PMC
April 2013

Inhibition of estradiol synthesis attenuates renal injury in male streptozotocin-induced diabetic rats.

Am J Physiol Renal Physiol 2011 Sep 8;301(3):F634-40. Epub 2011 Jun 8.

Department of Physiology and Biophysics, Univ. of Mississippi Medical Center, Jackson, 39216, USA.

We previously showed that the male streptozotocin (STZ)-induced diabetic rat exhibits decreased circulating testosterone and increased estradiol levels. While supplementation with dihydrotestosterone is partially renoprotective, the aim of the present study was to examine whether inhibition of estradiol synthesis, by blocking the aromatization of testosterone to estradiol using an aromatase inhibitor, can also prevent diabetes-associated renal injury. The study was performed on male Sprague-Dawley nondiabetic, STZ-induced diabetic, and STZ-induced diabetic rats treated with 0.15 mg/kg of anastrozole, an aromatase inhibitor (Da) for 12 wk. Treatment with anastrozole reduced diabetes-associated increases in plasma estradiol by 39% and increased plasma testosterone levels by 187%. Anastrozole treatment also attenuated urine albumin excretion by 42%, glomerulosclerosis by 30%, tubulointerstitial fibrosis by 32%, along with a decrease in the density of renal cortical CD68-positive cells by 50%, and protein expression of transforming growth factor-β by 20%, collagen type IV by 29%, tumor necrosis factor-α by 28%, and interleukin-6 by 25%. Anastrozole also increased podocin protein expression by 18%. We conclude that blocking estradiol synthesis in male STZ-induced diabetic rats is renoprotective.
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http://dx.doi.org/10.1152/ajprenal.00718.2010DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3174551PMC
September 2011

Sex differences in vasopressin V₂ receptor expression and vasopressin-induced antidiuresis.

Am J Physiol Renal Physiol 2011 Feb 1;300(2):F433-40. Epub 2010 Dec 1.

Division of Nephrology and Hypertension, Department of Medicine, Georgetown University Medical Center, Washington, DC 20007, USA.

The renal vasopressin V(2) receptor (V(2)R) plays a critical role in physiological and pathophysiological processes associated with arginine vasopressin (AVP)-induced antidiuresis. Because clinical data suggests that females may be more prone to hyponatremia from AVP-mediated antidiuresis, we investigated whether there are sex differences in the expression and function of the renal V(2)R. In normal Sprague-Dawley rat kidneys, V(2)R mRNA and protein expression was 2.6- and 1.7-fold higher, respectively, in females compared with males. To investigate the potential physiological implications of this sex difference, we studied changes in urine osmolality induced by the AVP V(2)R agonist desmopressin. In response to different doses of desmopressin, there was a graded increase in urine osmolality and decrease in urine volume during a 24-h infusion. Females showed greater mean increases in urine osmolality and greater mean decreases in urine volume at 0.5 and 5.0 ng/h infusion rates. We also studied renal escape from antidiuresis produced by water loading in rats infused with desmopressin (5.0 ng/h). After 5 days of water loading, urine osmolality of both female and male rats escaped to the same degree physiologically, but V(2)R mRNA and protein in female kidneys was reduced to a greater degree (-63% and -73%, respectively) than in males (-32% and -48%, respectively). By the end of the 5-day escape period, renal V(2)R mRNA and protein expression were reduced to the same relative levels in males and females, thereby abolishing the sex differences in V(2)R expression seen in the basal state. Our results demonstrate that female rats express significantly more V(2)R mRNA and protein in kidneys than males, and that this results physiologically in a greater sensitivity to V(2)R agonist administration. The potential pathophysiological implications of these results are that females may be more susceptible to the development of dilutional hyponatremia because of a greater sensitivity to endogenously secreted AVP.
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http://dx.doi.org/10.1152/ajprenal.00199.2010DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3044003PMC
February 2011

Accelerated fracture healing in mice lacking the 5-lipoxygenase gene.

Acta Orthop 2010 Dec 11;81(6):748-55. Epub 2010 Nov 11.

Department of Biochemistry and Molecular Biology, UMDNJ-New Jersey Medical School and Graduate School of Biological Sciences, Newark, USA.

Background And Purpose: Cyclooxygenase-2 (COX-2) promotes inflammation by synthesizing pro-inflammatory prostaglandins from arachidonic acid. Inflammation is an early response to bone fracture, and ablation of COX-2 activity impairs fracture healing. Arachidonic acid is also converted into leukotrienes by 5-lipoxygenase (5-LO). We hypothesized that 5-LO is a negative regulator of fracture healing and that in the absence of COX-2, excess leukotrienes synthesized by 5-LO will impair fracture healing.

Methods: Fracture healing was assessed in mice with a targeted 5-LO mutation (5-LO(KO) mice) and control mice by radiographic and histological observations, and measured by histomorphometry and torsional mechanical testing. To assess effects on arachidonic acid metabolism, prostaglandin E2, F2α, and leukotriene B4 levels were measured in the fracture calluses of control, 5-LO(KO) COX-1(KO), and COX-2(KO) mice by enzyme linked immunoassays.

Results: Femur fractures in 5-LO(KO) mice rapidly developed a cartilaginous callus that was replaced with bone to heal fractures faster than in control mice. Femurs from 5-LO(KO) mice had substantially better mechanical properties after 1 month of healing than did control mice. Callus leukotriene levels were 4-fold higher in mice homozygous for a targeted mutation in the COX-2 gene (COX-2(KO)), which indicated that arachidonic acid was shunted into the 5-LO pathway in the absence of COX-2.

Interpretation: These experiments show that 5-LO negatively regulates fracture healing and that shunting of arachidonic acid into the 5-LO pathway may account, at least in part, for the impaired fracture healing response observed in COX-2(KO) mice.
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http://dx.doi.org/10.3109/17453674.2010.533931DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3216088PMC
December 2010

Tumor necrosis factor-alpha antagonist etanercept decreases blood pressure and protects the kidney in a mouse model of systemic lupus erythematosus.

Hypertension 2010 Oct 9;56(4):643-9. Epub 2010 Aug 9.

Department of Physiology and Biophysics, Center for Excellence in Cardiovascular Renal Research, University of Mississippi Medical Center, 2500 North State St, Jackson, MS 39216-4505, USA.

Chronic inflammation has been implicated in the pathology of hypertension; however, the role for specific cytokines remains unclear. We tested whether tumor necrosis factor-α blockade with etanercept (Etan) reduces mean arterial pressure in a female mouse model of systemic lupus erythematosus (SLE). SLE is a chronic inflammatory disorder with prevalent hypertension. Thirty-week-old SLE (NZBWF1) and control mice (NZW/LacJ) received Etan (0.8 mg/kg SC weekly) for 4 weeks or vehicle. Mean arterial pressure (in millimeters of mercury) was increased in SLE mice (150±5 versus 113±5 in controls; P<0.05) and was lower in Etan-treated SLE mice (132±3) but not controls (117±5). Albuminuria (in micrograms per milligram of creatinine) was elevated in SLE mice (28 742±9032 versus 1075±883; P<0.05) and was lower in Etan-treated SLE mice (8154±3899) but not control animals (783±226). Glomerulosclerosis (in percentage of glomeruli) was evident in SLE mice (2.5±1.6 versus 0.0±0.0 in controls; P<0.05) and was ameliorated in Etan-treated SLE mice (0.1±0.1). Renal cortex CD68(+) cell staining (in percentage of area) was elevated in SLE mice (4.75±0.80 versus 0.79±0.12 in controls; P<0.05) and was lower in Etan-treated SLE mice (2.28±0.32) but not controls (1.43±0.25). Renal cortex NADPH oxidase activity (relative light units per milligram of protein) was higher in SLE mice compared with controls (10 718±1276 versus 7584±229; P<0.05) and lowered in Etan-treated SLE mice (6645±490). Renal cortex nuclear factor κB (phosphorylated and nonphosphorylated) was increased in SLE mice compared with controls and lower in Etan-treated SLE mice. These data suggest that TNF-α mechanistically contributes to the development of hypertension in a chronic inflammatory disease through increased renal nuclear factor κB, oxidative stress, and inflammation.
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http://dx.doi.org/10.1161/HYPERTENSIONAHA.110.157685DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2989495PMC
October 2010

Expression of aromatase, androgen and estrogen receptors in peripheral target tissues in diabetes.

Steroids 2010 Nov 11;75(11):779-87. Epub 2010 Jan 11.

Department of Medicine, Georgetown University Medical Center, 4000 Reservoir Road NW, Washington, DC 20057, United States.

Our previous studies have shown that diabetes in the male streptozotocin (STZ)-induced diabetic rat is characterized by a decrease in circulating testosterone and concomitant increase in estradiol levels. Interestingly, this increase in estradiol levels persists even after castration, suggesting extra-testicular origins of estradiol in diabetes. The aim of the present study was to examine whether other target organs of diabetes may be sources of estradiol. The study was performed in male Sprague-Dawley non-diabetic (ND), STZ-induced diabetic (D) and STZ-induced diabetic castrated (Dcas) rats (n=8-9/group). 14 weeks of diabetes was associated with decreased testicular (ND, 26.3+/-4.19; D, 18.4+/-1.54; P<0.05), but increased renal (ND, 1.83+/-0.92; D, 7.85+/-1.38; P<0.05) and ocular (D, 23.4+/-3.66; D, 87.1+/-28.1; P<0.05) aromatase activity. This increase in renal (Dcas, 6.30+/-1.25) and ocular (Dcas, 62.7+/-11.9) aromatase activity persisted after castration. The diabetic kidney also had increased levels of tissue estrogen (ND, 0.31+/-0.01; D, 0.51+/-0.11; Dcas, 0.45+/-0.08) as well as estrogen receptor alpha protein expression (ND, 0.63+/-0.09; D, 1.62+/-0.28; Dcas, 1.38+/-0.20). These data suggest that in male STZ-induced diabetic rats, tissues other than the testis may become sources of estradiol. In particular, the diabetic kidney appears to produce estradiol following castration, a state that is associated with a high degree or renal injury. Overall, our data provides evidence for the extra-testicular source of estradiol that in males, through an intracrine mechanism, may contribute to the development and/or progression of end-organ damage associated with diabetes.
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http://dx.doi.org/10.1016/j.steroids.2009.12.012DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2891268PMC
November 2010

A comparison of the effects of ibuprofen and rofecoxib on rabbit fibula osteotomy healing.

Acta Orthop 2009 Oct;80(5):597-605

Department of Biochemistry, University of Medicine and Dentistry of New Jersey, New Jersey Medical School, Newark, NJ, USA.

Background And Purpose: Non-steroidal anti-inflammatory drugs (NSAIDs) inhibit cyclooxygenase (COX) activity, which is the rate-limiting enzyme in the synthesis of prostaglandins. Previous studies have indicated that NSAID therapy, and in particular NSAIDs that specifically target the inflammatory cyclooxygenase (COX-2), impair bone healing. We compared the effects of ibuprofen and rofecoxib on fibula osteotomy healing in rabbits to determine whether nominal, continuous inhibition of COX-2 with rofecoxib would differentially affect fracture healing more than cyclical inhibition of COX-2 using ibuprofen, which inhibits COX-1 and COX-2 and has a short half-life in vivo.

Methods: Bilateral fibula osteotomies were done in 67 skeletally mature male New Zealand white rabbits. The rabbits were treated with placebo, rofecoxib (12.5 mg once a day), or ibuprofen (50 mg 3 times a day) for 28 days after surgery. Plasma ibuprofen levels were measured by HPLC analysis. Bone healing was assessed by histomorphometry at 3 and 6 weeks after osteotomy, and at 6 and 12 weeks by torsional mechanical testing.

Results: Plasma ibuprofen levels peaked and declined between successive doses. Fracture callus morphology was abnormal in the rofecoxib-treated rabbits and torsional mechanical testing showed that fracture healing was impaired. Ibuprofen treatment caused persistence of cartilage within the fracture callus and reduced peak torque at 6 weeks after osteotomy as compared to the fibulas from the placebo-treated rabbits. In the specimens allowed to progress to possible healing, non-union was seen in 5 of the 26 fibulas from the rofecoxib-treated animals as compared to 1 of 24 in the placebo group and 1 of 30 in the ibuprofen treatment group.

Interpretation: Continuous COX-2 inhibition as modeled by rofecoxib treatment appears to be more deleterious to fracture repair than cyclical cyclooxygenase inhibition as modeled by ibuprofen treatment. Ibuprofen treatment appeared to delay bone healing based upon the persistence of cartilage within the fracture callus and diminished shear modulus. Despite the ibuprofen-induced delay, rofecoxib treatment produced worse fracture (osteotomy) healing than ibuprofen treatment.
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http://dx.doi.org/10.3109/17453670903316769DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2823333PMC
October 2009

Dose-dependent effects of dihydrotestosterone in the streptozotocin-induced diabetic rat kidney.

Am J Physiol Renal Physiol 2009 Aug 3;297(2):F307-15. Epub 2009 Jun 3.

Department of Medicine, Georgetown University Medical Center, Washington, District of Columbia, USA.

We recently reported that castration exacerbates albuminuria, glomerulosclerosis, and tubulointerstitial fibrosis associated with diabetic renal disease. The aim of the present study was to examine whether these effects of castration can be attenuated with dihydrotestosterone (DHT) supplementation. The study was performed in castrated male Sprague-Dawley, streptozotocin-induced diabetic rats treated with 0 mg/day DHT (DHT(0)), 0.75 mg/day DHT (DHT(0.75)), or 2.0 mg/day DHT (DHT(2.0)) for 14 wk. Treatment with 0.75 mg/day DHT attenuated castration-associated increases in urine albumin excretion (DHT(0), 81.2 +/- 18.1; DHT(0.75), 26.57 +/- 5.8 mg/day; P < 0.05), glomerulosclerosis (DHT(0), 1.1 +/- 0.79; DHT(0.75), 0.43 +/- 0.043 arbitrary units; P < 0.001), tubulointerstitial fibrosis (DHT(0), 1.3 +/- 0.12; DHT(0.75), 1.1 +/- 0.096 AU; P < 0.05), collagen type IV [DHT(0), 3.2 +/- 0.11; DHT(0.75), 2.1 +/- 0.070 relative optical density (ROD); P < 0.01], transforming growth factor-beta (DHT(0), 3.2 +/- 0.16; DHT(0.75), 2.1 +/- 0.060 ROD; P < 0.01), IL-6 (DHT(0), 0.37 +/- 0.011; DHT(0.75), 0.27 +/- 0.014 ROD; P < 0.05), and protein expression and reduced CD68-positive cell abundance (DHT(0), 17 +/- 0.86; DHT(0.75), 4.4 +/- 0.55 cells/mm(2); P < 0.001). In contrast, treatment with 2.0 mg/day DHT exacerbated all these parameters. These data suggest that the detrimental effects of castration in the diabetic kidney can be attenuated with low doses of DHT, whereas high doses augment the adverse effects of castration, and these effects appear to be influenced by estradiol. We conclude that the effects of DHT are dose dependent but caution should be taken when DHT supplementation is considered in the treatment of diabetic renal disease.
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http://dx.doi.org/10.1152/ajprenal.00135.2009DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2724246PMC
August 2009

Comparison of fracture healing among different inbred mouse strains.

Calcif Tissue Int 2008 Jun;82(6):465-74

Department of Biochemistry and Molecular Biology, UMDNJ-New Jersey Medical School and Graduate School of Biomedical Sciences, 185 South Orange Avenue, Newark, NJ 07103, USA.

Quantitative trait locus analysis can be used to identify genes critically involved in biological processes. No such analysis has been applied to identifying genes that control bone fracture healing. To determine the feasibility of such an approach, healing of femur fractures was measured between C57BL/6, DBA/2, and C3H inbred strains of mice. Healing was assessed by radiography and histology and measured by histomorphometry and biomechanical testing. In all strains, radiographic bridging of the fracture was apparent after 3 weeks of healing. Histology showed that healing occurred through endochondral ossification in all strains. Histomorphometric measurements found more bone in the C57BL/6 fracture calluses 7 and 10 days after fracture. In contrast, more cartilage was present after 7 days in the C3H callus, which rapidly declined to levels less than those of C57BL/6 or DBA/2 mice by 14 days after fracture. An endochondral ossification index was calculated by multiplying the callus percent cartilage and bone areas as a measure of endochondral ossification. At 7 and 10 days after fracture, this value was higher in C57BL/6 mice. Using torsional mechanical testing, normalized structural and material properties of the C57BL/6 healing femurs were higher than values from the DBA/2 or C3H mice 4 weeks after fracture. The data indicate that fracture healing proceeds more rapidly in C57BL/6 mice and demonstrate that genetic variability significantly contributes to the process of bone regeneration. Large enough differences exist between C57BL/6 and DBA/2 or C3H mice to permit a quantitative trait locus analysis to identify genes controlling bone regeneration.
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http://dx.doi.org/10.1007/s00223-008-9144-3DOI Listing
June 2008

Characterization of a closed femur fracture model in mice.

J Orthop Trauma 2004 Nov-Dec;18(10):687-95

Department of Orthopaedics, University of Medicine and Dentistry of New Jersey-New Jersey Medical School, Newark, NJ 07103, USA.

Objectives: The goal of this study was to develop and characterize a closed femur fracture model for mice that can be used for the molecular and genetic analysis of fracture healing.

Study Design: Longitudinal time study of species-specific fracture healing.

Methods: A protocol was developed for creating reproducible, closed femur fractures in mice. Impending fractures were stabilized by retrograde insertion of a 0.01-inch-diameter, stainless steel wire into the intramedullary canal. The intramedullary wire was held in place with a wedge made from the first 2 mm of a 30-gauge needle. Fractures were produced by 3-point bending. Fracture healing was assessed by radiography, histology, and torsional mechanical testing.

Results: The mouse femur fracture technique produced good results with minimal loss of animals. Of the 246 mice used in the study, 22 mice were excluded due to poor fracture quality (8), loss of fracture stabilization (6), or to anesthesia death (8). Radiography showed a consistent pattern of fracture healing between mice with peak fracture callus volume evident at 10 (15 mice) to 14 days (18 mice) after fracture. Fracture bridging was apparent in all 3-week postfracture radiographs (35 mice). Histologic examination of 117 specimens at 9 time points showed chondrocyte differentiation within the fracture callus by 7 days after fracture, endochondral ossification occurring by 10 days after fracture, and bone remodeling evident as early as 3 weeks after fracture. Despite radiologic and histologic evidence of fracture bridging after 3 weeks, torsional mechanical testing of 68 mice at 3, 4, 6, and 12 weeks after fracture (group size of 15 to 18 mice at each time point) indicated that significant increases in structural or material strength did not occur until 6 to 12 weeks after fracture.

Conclusions: Femur fracture healing in mice follows a typical endochondral ossification pathway with fracture bridging occurring approximately 1 week faster in mice than rats. This fracture model is amenable to the molecular and genetic analysis of fracture healing using different inbred, transgenic, and knockout strains of mice.
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http://dx.doi.org/10.1097/00005131-200411000-00006DOI Listing
February 2005