text:
Improvement of endothelial nitric oxide synthase activity retards the progression of diabetic nephropathy in db/db mice.
ABSTRACT Impaired endothelial nitric oxide synthase (eNOS) activity may be involved in the pathogenesis of diabetic nephropathy. To test this, we used the type 2 diabetic db/db mouse (BKS background) model and found impaired eNOS dimerization and phosphorylation along with moderate glomerular mesangial expansion and increased glomerular basement membrane (GBM) thickness at 34 weeks of age. Cultured murine glomerular endothelial cells exposed to high glucose had similar alterations in eNOS dimerization and phosphorylation. Treatment with sepiapterin, a stable precursor of the eNOS cofactor tetrahydrobiopterin, or the nitric oxide precursor L-arginine corrected changes in eNOS dimerization and phosphorylation, corrected permeability defects, and reduced apoptosis. Sepiapterin or L-arginine, administered to db/db mice from weeks 26 to 34, did not significantly alter hyperfiltration or affect mesangial expansion, but reduced albuminuria and GBM thickness, and decreased urinary isoprostane and nitrotyrosine excretion (markers of oxidative stress). Although there was no change in glomerular eNOS monomer expression, both sepiapterin and L-arginine partially reversed the defect in eNOS dimerization and phosphorylation. Hence, our results support an important role for eNOS dysfunction in diabetes and suggest that sepiapterin supplementation might have therapeutic potential in diabetic nephropathy.Kidney International advance online publication, 11 July 2012; doi:10.1038/ki.2012.248.
Article: Effects of Folic Acid on Renal Endothelial Function in Patients with Diabetic Nephropathy: Results from a Randomized Trial.[Show abstract] [Hide abstract]
ABSTRACT: Endothelial dysfunction has been shown to promote podocyte injury and albuminuria in diabetes, highlighting the importance of the interaction between renal endothelial cells and podocytes. Folic acid (FA) improves nitric oxide synthase (NOS) function and ameliorates progression of diabetic nephropathy in animal models. We tested whether high-dose FA treatment improves renal endothelial function and albuminuria in human subjects with incipient diabetic nephropathy. Following a double-blind, randomized cross-over design, 28 patients with type 2 diabetes and albuminuria were allocated to 4-weeks treatment with placebo and high-dose FA (5 mg/day). Renal NO production determined as response of renal plasma flow (RPF) to NOS inhibition with N(G)-monomethyl-L-arginine (4.25 mg/kg i.v.), renal oxidant stress as the response of RPF to vitamin C infusion (3mg/kg) and albuminuria were determined after each treatment phase. Neither the reduction of RPF to L-NMMA nor the increase of RPF to vitamin C infusion differed between treatment phases (ΔRPF to L-NMMA: -74±71 ml/min/m2 during placebo versus -63±56 ml/min/m2 during FA, P=0.57; ΔRPF to vitamin C: +93±118 ml/min/m2 versus +94±108 ml/min/m2; P=0.70). In line with the lack of effect on the renal endothelium, albuminuria was not affected by FA treatment (110±179 mg/day during placebo versus 87±146 mg/day during FA; P=0.12). High-dose FA treatment does not improve renal endothelial function and fails to reduce albuminuria in human subjects with diabetic nephropathy. Novel treatment options for oxidant stress and endothelial dysfunction in patients with diabetes are urgently needed.Clinical Science 04/2014; · 4.86 Impact Factor
Article: Diffuse glomerular nodular lesions in diabetic pigs carrying a dominant-negative mutant hepatocyte nuclear factor 1-alpha, an inheritant diabetic gene in humans.[Show abstract] [Hide abstract]
ABSTRACT: Glomerular nodular lesions, known as Kimmelstiel-Wilson nodules, are a pathological hallmark of progressive human diabetic nephropathy. We have induced severe diabetes in pigs carrying a dominant-negative mutant hepatocyte nuclear factor 1-alpha (HNF1α) P291fsinsC, a maturity-onset diabetes of the young type-3 (MODY3) gene in humans. In this model, glomerular pathology revealed that formation of diffuse glomerular nodules commenced as young as 1 month of age and increased in size and incidence until the age of 10 months, the end of the study period. Immunohistochemistry showed that the nodules consisted of various collagen types (I, III, IV, V and VI) with advanced glycation end-product (AGE) and Nε-carboxymethyl-lysine (CML) deposition, similar to those in human diabetic nodules, except for collagen type I. Transforming growth factor-beta (TGF-β) was also expressed exclusively in the nodules. The ultrastructure of the nodules comprised predominant interstitial-type collagen deposition arising from the mesangial matrices. Curiously, these nodules were found predominantly in the deep cortex. However, diabetic pigs failed to show any of the features characteristic of human diabetic nephropathy; e.g., proteinuria, glomerular basement membrane thickening, exudative lesions, mesangiolysis, tubular atrophy, interstitial fibrosis, and vascular hyalinosis. The pigs showed only Armanni-Ebstein lesions, a characteristic tubular manifestation in human diabetes. RT-PCR analysis showed that glomeruli in wild-type pigs did not express endogenous HNF1α and HNF1β, indicating that mutant HNF1α did not directly contribute to glomerular nodular formation in diabetic pigs. In conclusion, pigs harboring the dominant-negative mutant human MODY3 gene showed reproducible and distinct glomerular nodules, possibly due to AGE- and CML-based collagen accumulation. Although the pathology differed in several respects from that of human glomerular nodular lesions, the somewhat acute and constitutive formation of nodules in this mammalian model might provide information facilitating identification of the principal mechanism underlying diabetic nodular sclerosis.PLoS ONE 01/2014; 9(3):e92219. · 3.53 Impact Factor
Article: Vascular Endothelial Growth Factor-Receptor 1 Inhibition Aggravates Diabetic Nephropathy through eNOS Signaling Pathway in db/db Mice.[Show abstract] [Hide abstract]
ABSTRACT: The manipulation of vascular endothelial growth factor (VEGF)-receptors (VEGFRs) in diabetic nephropathy is as controversial as issue as ever. It is known to be VEGF-A and VEGFR2 that regulate most of the cellular actions of VEGF in experimental diabetic nephropathy. On the other hand, such factors as VEGF-A, -B and placenta growth factor bind to VEGFR1 with high affinity. Such notion instigated us to investigate on whether selective VEGFR1 inhibition with GNQWFI hexamer aggravates the progression of diabetic nephropathy in db/db mice. While diabetes suppressed VEGFR1, it did increase VEGFR2 expressions in the glomerulus. Db/db mice with VEGFR1 inhibition showed more prominent features with respect to, albuminuria, mesangial matrix expansion, inflammatory cell infiltration and greater numbers of apoptotic cells in the glomerulus, and oxidative stress than that of control db/db mice. All these changes were related to the suppression of diabetes-induced increases in PI3K activity and Akt phosphorylation as well as the aggravation of endothelial dysfunction associated with the inactivation of FoxO3a and eNOS-NOx. In cultured human glomerular endothelial cells (HGECs), high-glucose media with VEGFR1 inhibition induced more apoptotic cells and oxidative stress than did high-glucose media alone, which were associated with the suppression of PI3K-Akt phosphorylation, independently of the activation of AMP-activated protein kinase, and inactivation of FoxO3a and eNOS-NOx pathway. In addition, transfection with VEGFR1 siRNA in HGECs also suppressed PI3K-Akt-eNOS signaling. In conclusion, the specific blockade of VEGFR1 with GNQWFI caused severe renal injury related to profound suppression of the PI3K-Akt, FoxO3a and eNOS-NOx pathway, giving rise to the oxidative stress-induced apoptosis of glomerular cells in type 2 diabetic nephropathy.PLoS ONE 01/2014; 9(4):e94540. · 3.53 Impact Factor
Page 1
see commentary on page 1151
Improvement of endothelial nitric oxide synthase
activity retards the progression of diabetic
nephropathy in db/db mice
Huifang Cheng1, Hanmin Wang1, Xiaofeng Fan1, Paisit Paueksakon2and Raymond C. Harris1
1Division of Nephrology, Department of Medicine, George M. O'Brien Kidney and Urologic Diseases Center, Vanderbilt University School
of Medicine, Nashville Veterans Affairs Hospital, Nashville, Tennessee, USA and2Department of Pathology, Microbiology and
Immunology, George M. O'Brien Kidney and Urologic Diseases Center, Vanderbilt University School of Medicine, Nashville Veterans
Affairs Hospital, Nashville, Tennessee, USA
Impaired endothelial nitric oxide synthase (eNOS) activity
may be involved in the pathogenesis of diabetic
nephropathy. To test this, we used the type 2 diabetic db/db
mouse (BKS background) model and found impaired eNOS
dimerization and phosphorylation along with moderate
glomerular mesangial expansion and increased glomerular
basement membrane (GBM) thickness at 34 weeks of age.
Cultured murine glomerular endothelial cells exposed to high
glucose had similar alterations in eNOS dimerization and
phosphorylation. Treatment with sepiapterin, a stable
precursor of the eNOS cofactor tetrahydrobiopterin, or the
nitric oxide precursor L-arginine corrected changes in eNOS
dimerization and phosphorylation, corrected permeability
defects, and reduced apoptosis. Sepiapterin or L-arginine,
administered to db/db mice from weeks 26 to 34, did not
significantly alter hyperfiltration or affect mesangial
expansion, but reduced albuminuria and GBM thickness, and
decreased urinary isoprostane and nitrotyrosine excretion
(markers of oxidative stress). Although there was no change
in glomerular eNOS monomer expression, both sepiapterin
and L-arginine partially reversed the defect in eNOS
dimerization and phosphorylation. Hence, our results
support an important role for eNOS dysfunction in diabetes
and suggest that sepiapterin supplementation might have
therapeutic potential in diabetic nephropathy.
Kidney International (2012) 82, 1176–1183; doi:10.1038/ki.2012.248;
published online 11 July 2012
endothelial cells; sepiapterin
Multiple mechanisms contribute to the development of
diabetic nephropathy, and there is increasing evidence that all
three types of glomerular cells are potential targets for
Specifically, there is increasing evidence that
dysfunction of the fenestrated glomerular endothelial cells
(GEnCs) is an important contributory factor to progressive
diabetic glomerular injury.2–5Nitric oxide (NO) is an
important endothelium-derived mediator, with multiple
actions, including vasodilation and antiplatelet, antiproli-
ferative, antiadhesive, permeability-decreasing, and anti-
inflammatory properties.3Previous studies have examined
whether there are alterations in NO production and/or
bioavailability in diabetes but have provided conflicting
results. In vitro studies have shown that high glucose (HG)
medium could either induce4,5or inhibit6endothelial nitric
oxide synthase (eNOS) expression in cultured endothelial
cells. In animal model of diabetes mellitus, eNOS gene
expression has been described to be either unchanged,7
diminished,8,9or increased.10,11Hohenstein et al.12recently
reported upregulated eNOS expression in glomeruli from
type 2 diabetic patients with nephropathy. In contrast, we
and others have found that eNOS insufficiency will accelerate
nephropathy in mouse models of both type 1 and type 2
diabetes.13–19Therefore, the underlying mechanisms and
potential roles of impaired eNOS in glomerular endothelium
during the development of diabetic nephropathy still remain
incompletely understood.14
HG induced GEnC impairment and eNOS dysfunction
Pilot studies indicated that incubation of cultured mouse
GEnCs with HG for 48h demonstrated significant alterations.
At that time, GEnCs in HG increased bovine serum albumin
(BSA) clearance compared with normal glucose (NG) medium
or mannitol (9.9±0.6% in NG and 11.0±1.0% in mannitol vs.
40.8±1.7% in HG after 40min, n¼5, Po0.05) (Figure 1a)
without significantly altering proliferation (data not shown).
In addition, HG induced profound apoptosis in GEnCs
(1±0.2% in NG vs. 7.9±0.4% in HG, n¼4, Po0.05)
original article
& 2012 International Society of Nephrology
Received 7 February 2012; revised 20 April 2012; accepted 22 May 2012;
published online 11 July 2012
Correspondence: Raymond C. Harris, Division of Nephrology, Department
of Medicine, C3121 MCN, George M. O'Brien Kidney and Urologic Diseases
Center, Vanderbilt University School of Medicine, Nashville Veterans Affairs
Hospital, Nashville, Tennessee 37232, USA. E-mail: email@example.com
Kidney International (2012) 82, 1176–1183
Page 2
(Figure 1b and c). Incubation in the high-glucose medium
decreased eNOS dimerization (Figure 2a) and phosphoryla-
tion at Ser1179 (Figure 2b) without changing eNOS
monomer expression or phosphorylation at Thr479, whereas
incubation with mannitol as an osmolar control did not have
the same effects (Supplementary Figure S1a and b online).
HG also decreased NOS activity, indicated by decreased levels
of the stable end products of NO (nitrate/nitrite) (to
0.53±0.07 fold of NG. n¼4, Po0.05 compared with NG
or mannitol control) (Figure 2c and Supplementary Figure
S1c online).
Effects of sepiapterin and L-arginine on HG–induced GEnC
We treated the GEnCs exposed to high-glucose medium with
the NO precursor, L-arginine, or the tetrahydrobiopterin
(BH4) cofactor, sepiapterin. Either L-arginine or sepiapterin
partially corrected the HG–induced defects in eNOS dimer-
ization and phosphorylation (Figure 2a and b), as well as
eNOS activity (to 0.86±0.05- and 0.83±0.08-fold control
by L-arginine or sepiapterin, respectively, n¼4, Po0.05
compared with untreated) (Figure 2c). In addition, both
L-arginine and sepiapterin significantly blunted GenC altera-
tions in permeability and apoptosis induced by incubation in
the high-glucose medium (Figure 1b–d).
eNOS dysfunction in db/db mice
db/db mice are a well-established type 2 diabetic model, but
some strains are relatively resistant to diabetic renal injury
despite the significant hyperglycemia.15,16db/db mice on the
BKS background have been shown to develop hyperglycemia
and diabetic renal injury.17,18We found that at 34 weeks, db/
db (BKS) mice had developed glomerulopathy characterized
by mesangial matrix accumulation (Figure 3a), foot process
effacement, and increased glomerular basement membrane
(GBM) thickness (154±6 in BKS control vs. 335±23nm in
untreated db/db mice) (Figure 3a and b) and had significant
albuminuria (alb/cre: 18±3 in BKS control vs. 759±281 in
untreated db/db, n¼8, Po0.05) (Figure 3c) and hyperfiltra-
tion (glomerular filtration rate: 206±33ml/min in BKS
control; 403±36 in untreated db/db, n¼6–12, Po0.05)
(Figure 3d). Furthermore, urine F2-isoprostanes, a marker of
oxidative stress, increased in the diabetic mice (5.7±0.9 in
db/db mice vs. 1.4±0.1ng 8-iso-PGF2a/mg Cr in BKS
102030 4050
Total (PI) Apoptosis (TUNEL)
BSA clearance (%)
BSA clearance (%)
Apoptosis (%)
Figure 1|High glucose increased bovine serum albumin (BSA) permeability and apoptosis in glomerular endothelial cells (GEnCs);
these effects were inhibited by sepiapterin or L-arginine. (a) High glucose (HG), but not mannitol, increased permeability in GEnCs,
indicated by BSA clearance at 5–40min. *Po0.05 HG versus normal medium (NG) or osmolality control (mannitol at same concentration).
(b) Sepiapterin (Sep) and L-arginine (L-arg) attenuated HG increases in permeability, *Po0.05 HG versus normal medium or HG with Sep or
L-arg coincubation. (c) Effect of HG on apoptosis in GEnCs. Representative images of terminal transferase dUTP nick-end labeling (TUNEL)
assay from four independent experiments. (d) Sep and L-arg prevented HG-induced apoptosis. *Po0.05 HG versus normal medium or
HG with Sep or L-arg coincubation.
Kidney International (2012) 82, 1176–1183
H Cheng et al.: Sepiapterin protects against glomerular injury
original article
Page 3
control, n¼6–11, Po0.05) (Figure 3e). Increased oxidative
stress in db/db mice was also confirmed by the accumulation
of nitrotyrosine, another superoxide marker (Figure 3f).
To investigate the potential role of eNOS in the diabetic
renal injury, we determined glomerular eNOS expression.
There were no differences in eNOS monomer expression
between control and diabetic mice (Figure 4a), which was
further confirmed by immunohistochemical staining (data
not shown). However, glomerular eNOS dimerization
(Figure 4b) and phosphorylation at Ser1179 (Figure 4c)
decreased by B50% in db/db mice, whereas phosphorylation
at Thr497 was unchanged (Figure 4c). HG–mediated
alterations in endothelial function have been reported to be
associated with reduced intracellular BH4.19,20In agreement
with these findings, we found that glomerular BH4in db/db
mice (0.7±0.1ng/mg pro.) was significantly lower than BKS
control (2.3±0.2ng/mg pro., n¼6, Po0.05).
Sepiaterin or L-arginine partially restored eNOS function and
attenuated renal injury in db/db mice
To examine the impact of L-arginine or sepiapterin in vivo, we
administered these agents to 26-week-old db/db mice for 8
weeks. Sepiapterin treatment significantly restored glomeru-
lar BH4 levels (to 1.9±0.3ng/mg pro., n¼6, Po0.05
compared with untreated db/db). Consistent with the in
vitro results, both
L-arginine and sepiapterin improved
glomerular eNOS function in db/db mice, indicated by
restoration of eNOS dimerization (Figure 4b) and phosphor-
ylation at Ser1179 (Figure 4c). In addition, oxidative stress,
indicated by nitrotyrosine accumulation and urinary F2-
isoprostane levels, was significantly decreased (3.1±0.2ng
8-iso-PGF2a/mg Cr for L-arginine and 2.7±0.2 for sepiap-
terin, n¼6–11, Po0.05 compared with untreated) (Figure 3e
and f).
Neither L-arginine nor sepiapterin treatment markedly
affected the increased mesangial expansion, but both of
them reduced GBM thickness and foot-process effacement
in db/db mice (Figure 3a and b). They also significantly
reduced albuminuria (sepiapterin: 40±7 and
163±37mgalb/mg Cr, respectively, n¼6–8, Po0.05 vs. un-
treated db/db) (Figure 3c) without affecting glomerular filtration
rate (398±16 in sepiapterin and 425±50ml/min in L-arginine,
respectively, n¼6, Po0.05 compared with BKS control; not
significantly different vs. untreated db/db) (Figure 3d).
Our studies showed that HG induced dysfunction of GEnCs
in vitro, as indicated by the increased permeability to BSA
and apoptosis, as well as decreased eNOS dimerization,
impaired eNOS phosphorylation at Ser 1179, and decreased
NO production. All of these abnormalities were ameliorated
by coincubation with either the NO precursor, L-arginine, or
with sepiapterin, a stable precursor of the eNOS cofactor,
tetrahydrobiopterin. These in vitro findings were confirmed
in db/db (BKS) mice, in which supplementation with either
sepiapterin or L-arginine attenuated albuminuria and GBM
thickness and decreased oxidative stress.
NO has an important role in controlling vascular tone and
modulating platelet activation. Decreased NO bioactivity
contributes to arterial thrombosis and endothelial dysfunc-
tion.21Chronic kidney diseases, including diabetic nephro-
pathy, are often characterized by relative NO deficiency,
eNOS dimer
Ser 1179
Thr 497
NOS activity (fold of NG)
Figure 2|Sepapterin (Sep) and L-arginine (L-arg) reversed high glucose–induced endothelial nitric oxide synthase (eNOS)
impairment in glomerular endothelial cells (GEnCs). (a) High glucose did not alter eNOS expression, but decreased eNOS dimerization in
GEnCs, which could be prevented by Sep or L-arg. Representative photo from three independent experiments. (b) High glucose decreased
eNOS phosphorylation at Ser1179, without significant change at Thr479. Sep or L-arg coincubation reversed phosphorylation at Ser1179.
Representative photo from three independent experiments. (c) High glucose reduced nitrate/nitrite production by GEnCs, which was
corrected by supplementation with Sep or L-arg. *Po0.05 compared with normal medium or high glucose with Sep or L-arg. HG, high
glucose; NG, normal glucose.
Kidney International (2012) 82, 1176–1183
original article
H Cheng et al.: Sepiapterin protects against glomerular injury
Page 4
which may contribute to disease progression.22NO is
synthesized from L-arginine by its synthases. eNOS is the
major source of NO production in vascular endothelial cells,
including the endothelium of preglomerular vessels and
Accelerated glomerular injury in diabetic mice with eNOS
deficiency strongly suggests that deficient eNOS activity may
have an important role in the pathogenesis of diabetic
nephropathy,13,24,25although there have been conflicting
results about eNOS regulation in the diabetic kidney, which
may relate to whether studies were conducted in early or
late injury.7–12,26Previous studies in isolated blood vessels27,28
and cultured umbilical vein cells29,30have been used as in vitro
systems to examine the role of eNOS on endothelial function.
Notably, GEnCs represent a microvascular endothelial cell
with unique characteristics31,32and have a critical role in
the pathogenesis of diabetic nephropathy. In the current
study, we focused on the impact of hyperglycemia on
eNOS impairment in cultured mouse GEnCs. In cultured
GEnCs, we found that HG increased both permeability to
BKS db/db controldb/db+Sepdb/db+L-arg
Albumin (μg)/creatinine
GFR (μl/min)
GBM thickness (nm)
db/db control
db/db control
db/db control
db/db control
BKS db/db control db/db+Sep db/db+L-arg
Figure 3|Sepiapterin (Sep) and L-arginine (L-arg) attenuated renal injury in db/db mice. (a) The upper panel indicates glomerular
mesangial expansion in db/db mice at 34 weeks by Periodic Acid Schiff staining, Bars: 20mm; the lower panel indicates glomerular basement
membrane (GBM) thickness and foot process effacement (red arrow) in db/db mice with or without treatment with Sep/L-arg, bars: 2mm.
(b) GBM thickness. *Po0.05, compared with wild-type BKS;#Po0.05, versus each treatment. (c) Sep or L-arg decreased albuminuria in
db/db mice at 34 weeks of age. *Po0.05, compared with wild-type BKS;#Po0.05, versus each treatment. (d) Sep or L-arg had no effect
on hyperfiltration seen in db/db mice, determined by fluorescein isothiocyanate-inulin clearance. *Po0.05, BKS versus db/db mice regardless
of treatments. (e) Sep or L-arg decreased elevations in urine isoprostane excretion in db/db mice. *Po0.05, compared with wild-type BKS;
#Po0.05, versus each treatment. (f) Increased nitrotyrosine accumulation in db/db mice was attenuated by Sep or L-arg treatment.
Representative photo from three independent experiments. GFR, glomerular filtration rate.
Kidney International (2012) 82, 1176–1183
H Cheng et al.: Sepiapterin protects against glomerular injury
original article
Page 5
BSA and apoptosis, in association with endothelial eNOS
There is evidence that activation of eNOS requires
formation of a homodimer through a linkage between the
N-terminal oxygenase domains.33Although we did not find
any significant alteration in eNOS monomer expression in
glomeruli from db/db mice at 34 weeks or in GEnCs in
response to HG, there was decreased eNOS dimerization.
We also found that in both diabetic glomeruli and high-
glucose GEnCs, eNOS phosphorylation was impaired at
Ser1179 without significant alteration at Thr497. A crucial
role for posttranslational control of eNOS bioactivity has been
suggested, owing to its long half-life at baseline (10–35h).34,35
Akt-dependent phosphorylation of eNOS at Ser1179 is
essential for endothelium-dependent relaxation,36whereas
phosphorylation at Thr497 is considered inhibitory.37
Reduced phosphorylation of eNOS at Ser1179 has previously
been reported in moderately hyperglycemic diabetic rats.38
In addition to decreased eNOS expression and decreased
eNOS activity due to uncoupling by reactive oxygen
species,22alterations in the availability of substrate and/or
essential cofactors may lead to decreased NO production.
When either the essential substrate,
essential cofactor, BH4, is limited, electron transfer from
eNOS flavins becomes uncoupled from L-arginine oxidation,
and superoxide is produced from the oxygenase domain,39–41
which may further reduce NO bioactivity and increase oxida-
tive stress within endothelial cells by scavenging NO and
forming peroxynitrite.42Reduced levels of both L-arginine
and BH4have been observed in diabetic rats,20,43with more
profound depletion in endothelial cells than in plasma.20
Modulation of the arginine–NO pathway by supplementa-
tion with arginine may have protective effects on diabetic
rats44and patients.20In a previous study, administration
of BH4to db/db mice did not affect glucose, triglyceride,
and cholesterol levels or body weight, but enhanced the
reactivity of small mesenteric arteries to phenylephrine.45
Acute intra-arterial infusion of BH4 leads to short-term
L-arginine, or the
improvements of forearm blood flow responses to endothelium-
dependent vasodilators in patients with type 2 diabetes.46
Sepiapterin, the immediate precursor of BH4via the biopterin
salvage pathway, is less sensitive to oxidative stress than BH4.47
It is noteworthy that administration of either L-arginine or
sepiapterin not only corrected the defects in eNOS dimeriza-
tion and phosphorylation but also decreased oxidative stress
and partially corrected functional and structural glomerular
abnormalities. Previous studies have also reported beneficial
effects of L-arginine on renal dilatation in animal models48and
humans.49The current in vitro study provided evidence for
a direct role of both sepiapterin and arginine on GEnCs,
independent of vasodilation.
In summary, we found that providing either a precursor for
NO production or a stable precursor of tetrahydrobiopterin
significantly improved the function of cultured GEnCs and
decreased progression of glomerular injury in a model of type 2
diabetes. Recently, a more stable preparation of oral BH4,
sapropterin dihydrochloride (Kuvan, BioMarin, Tiburon, CA),
has been developed and approved by the Food and Drug
Administration for treatment of patients with phenylketo-
nuria.50Our results provide evidence that BH4supplementa-
tion may have therapeutic potential in diabetic nephropathy.
Sepiapterin, eNOS polyclonal antiserum, and Nitrate/Nitrite Colori-
metric Assay Kits were from Cayman Chemical Co (Ann Arbor, MI).
Rabbit anti-phospho-serine-1179 eNOS (bovine) and anti-phospho-
Thr-497 polyclonal antibody were from Cell Signaling Technology
(Danvers, MA); anti-nitrotyrosine antibody was from Santa Cruz
Biotechnology (Santa Cruz, CA). The TUNEL (terminal transferase
dUTP nick-end labeling) apoptosis detection kit was from Upstate
(Lake Placid NY); other reagents were purchased from Sigma
Chemical (St Louis, MO).
Experimental animals
We used db/db mice on the nephropathy-susceptible BKS back-
ground51,52as a model of type 2 diabetes. Age-matched (26 weeks old)
eNOS dimer
Ser 1179
Thr 497
eNOS/β-Actin (fold control)
db/db control
db/db control
db/db control
eNOS dimer/monomer 0.8
Figure 4|Sepiapterin (Sep) and L-arginine (L-arg) reversed endothelial nitric oxide synthase (eNOS) impairment in db/db mice.
(a) No significant differences in glomerular eNOS expression were detected between BKS and db/db mice with or without treatment.
(n¼4; NS). (b) There was decreased eNOS dimerization in glomeruli from db/db mice, which was corrected by administration of Sep or
L-arg (n¼4, *Po0.05, compared with wild-type or treated groups). (c) There was decreased eNOS phosphorylation at Ser1179, but not at
Thr479, in glomeruli from db/db mice and normalization by Sep/L-arg treatment. Representative photo was from three independent
Kidney International (2012) 82, 1176–1183
original article
H Cheng et al.: Sepiapterin protects against glomerular injury
Page 6
male mice were divided into the following groups: (1) Wild type
(BKS); (2) db/db untreated (db/db control); (3) db/db mice treated
with sepiapterin, in which the mice were administered fresh
sepiapterin solution (10mg/kg/day by gavage) for 8 weeks (from
26 to 34 weeks); (4) db/db mice with L-arginine, in which L-arginine
(100mg/kg/day in the drinking water) was administered for the
same period. Mice were killed at the end of 34 weeks. All animal
procedures were approved by the Animal Care and Use Committee
of Vanderbilt University Medical Center.
Assessment of glomerular injury
Renal injury was assessed histologically with light microscopy by
hematoxylin and eosin, Periodic Acid Schiff, and Jones staining and
by electron microscopy. GBM thickness was randomly measured as
the distance between the endothelial cell and epithelial cell plasma
membranes ?20 for each sample and evaluated by the renal
pathologist. Urinary albumin levels were determined by enzyme-
linked immunosorbent assay (ELISA) using a murine micro-
albuminuria ELISA kit, AlbuwellM (Exocell, Philadelphia, PA).
The urine creatinine concentration was measured with a microplate
assay kit, Creatinine Companion (Exocell). All measurements were
recorded in duplicate, and albuminuria was determined as the ratio
of urinary albumin (mg/ml) to creatinine (mg/ml). Renal function
was evaluated by glomerular filtration rate as described previously.13
F2-isoprostane assay
F2-isoprostanes were measured in urine using stable isotope dilution
methods and gas chromatography negative ion chemical ionization
mass spectrometry,53,54and expressed as ng/mg urine creatinine.
Isolation of glomeruli
Glomeruli were isolated immediately after killing, using the
Dynabeads method modified from Takemoto et al.55Briefly, mice
were anesthetized by an injection of nembutal (0.05mg/g BW) and
perfused through the descending aorta with 8?107Dynabeads
diluted in phosphate-buffered saline. The kidneys were removed,
minced into 1-mm3pieces, and digested in collagenase (1mg/ml
collagenase A, 100U/ml deoxyribonuclease I in Hank's Balanced Salt
Solution (HBSS)) at 371C for 30min with gentle agitation. The
collagenase-digested tissue was gently pressed through a 100-mm cell
strainer using a flattened pestle, and the cell strainer was then
washed with 5ml of cold HBSS. The filtered cells were passed
through a new cell strainer without pressing and the cell strainer
washed with 5ml of cold HBSS. The cell suspension was then
centrifuged at 200g for 5min at 41C. The supernatant was discarded
and the cell pellet was resuspended in 2ml of cold HBSS. Finally,
glomeruli containing Dynabeads were gathered by a magnetic
particle concentrator and washed three times with cold HBSS.
During the procedure, kidney tissues were kept at 41C, except for
the collagenase digestion, which was kept at 371C. The preparation
consisted of 490% glomeruli.
Cultured mouse GEnCs
We used early-passage GEnCs from Michael Madaio,56grown in
DMEM/F12 medium with 10% FBS, at 371C in humidified air with
5% CO2. After quiescence for 16–24h, subsets of cells were exposed
to high-glucose (30mM) medium or osmolality control medium
(30mM mannitol) for 48h before experimentation. Subsets of cells
grown in high-glucose medium were coincubated with L-arginine
(3mM) or sepiapterin (100mM).57
Cultured cells or isolated glomeruli were homogenized as described
previously.58Proteins were resuspended in SDS sample buffer, diluted
in SDS buffer containing 2-mercaptoethanol (Sigma Chemical), and
boiled for 10min before loading. The samples were run on 8% sodium
dodecyl sulfate polyacrylamide gel electrophoresis gels under reducing
conditions and transferred onto polyvinylidene fluoride membrane
(Immobilion-P; Millipore, Bedford, MA). After blocking with 5%
nonfat milk in Tris-buffered saline, the membranes were exposed to
the primary antibody overnight at 41C, followed by horseradish
peroxidase–conjugated secondary antibodies. The horseradish perox-
idase signal was enhanced using the enhanced chemiluminescence
method, and the images were developed on high-performance
autoradiography film, Hyperfilm MP (Amersham Biosciences, Buck-
inghamshire, UK). Membranes were rehybridized with goat anti-b-
actin antibody (Santa Cruz Biotechnology) to normalize protein
loading. For detection of eNOS dimerization, we used nondenaturing,
low-temperature sodium dodecyl sulfate polyacrylamide gel electro-
phoresis to run gels with 50mg protein loading in each well without
boiling, and calculated eNOS dimer/monomer ratios.
Apoptosis detection
ATUNEL apoptosis detection kit (Upstate) was used for measurements.
Propidium iodide was used for counterstaining. The percentage of
apoptotic cells in 400 total cells from the same field was determined
for quantification.
Glomerular BH4(tetrahydrobiopterin)
Glomerular BH4was evaluated with the Tetrahydrobiopterin ELISA
Kit (MyBioSource, LLC, San Diego, CA) using the company's
instructions. BH4was normalized to protein content, expressing as
ng/mg pro.
Endothelial cell permeability
Permeability was detected by clearance of albumin across endo-
thelial monolayers as described previously.59,60Briefly, monolayers
were grown to confluence on the inserts of Transwell (3.0mm;
Becton Dickinson Labware, Franklin Lake, NJ, 8?105cells/insert).
After washing, the monolayers were covered immediately with
HBSS containing a trypan blue–albumin complex (trypan blue
0.035% and BSA 0.8%) and incubated at 371C with continuous
agitation. Samples were collected every 5min from the bottom well
up to 40min and were replaced by an equal volume of HBSS.
Absorbance at 590nm was measured from both the insert and the
lower well fluids. Results were expressed as clearance of BSA,
i.e., the amount of TB-BSA detected in the lower chamber, expressed
as a percentage of the total TB-BSA initially added to the upper
NOS activity from cultured GEnCs
Activity was measured with a Nitrate/Nitrite Colorimetric Assay Kit
(Cayman Chemical) from stable end products of the reaction of NO
with molecular oxygen, according to the manufacturer's instruc-
tions, normalized by the protein content and expressed as fold
control (cells gown in normal medium).
Statistical analyses
All values are presented as mean±s.e.m. ANOVA and Bonferroni
t-tests were used for statistical analysis, and differences were
considered significant when Po0.05.
Kidney International (2012) 82, 1176–1183
H Cheng et al.: Sepiapterin protects against glomerular injury
original article
Page 7
The authors declare no conflict of interest.
This work was supported by funds from the Department of Veterans
Affairs, the Vanderbilt George O'Brien Kidney and Urologic Diseases
Center (DK DK79341), DK51265, DK62794, and Vanderbilt Diabetes
Research and Training Center (DK20593).
Figure S1. High glucose induced eNOS dysfunction independent of
its osmolality.
Supplementary material is linked to the online version of the paper at
1.Nawroth PP, Isermann B. Mechanisms of diabetic nephropathy—old
buddies and newcomers part 1. Exp Clin Endocrinol Diab 2010; 118:
2.Patrakka J, Tryggvason K. Molecular make-up of the glomerular filtration
barrier. Biochem Biophys Res Commun 2010; 396: 164–169.
3.Stehouwer CD. Endothelial dysfunction in diabetic nephropathy: state of
the art and potential significance for non-diabetic renal disease. Nephrol
Dial Transplant 2004; 19: 778–781.
4.Hoshiyama M, Li B, Yao J et al. Effect of high glucose on nitric oxide
production and endothelial nitric oxide synthase protein expression in
human glomerular endothelial cells. Nephron Exp Nephrol 2003; 95:
5.Komers R, Allen TJ, Cooper ME. Role of endothelium-derived nitric oxide
in the pathogenesis of the renal hemodynamic changes of experimental
diabetes. Diabetes 1994; 43: 1190–1197.
6. Mohan S, Konopinski R, Yan B et al. High glucose-induced IKK-Hsp-90
interaction contributes to endothelial dysfunction. Am J Physiol Cell
Physiol 2009; 296: C182–C192.
7.Janssens S, Flaherty D, Nong Z et al. Human endothelial nitric oxide
synthase gene transfer inhibits vascular smooth muscle cell proliferation
and neointima formation after balloon injury in rats. Circulation 1998; 97:
8.Ju H, Zou R, Venema VJ et al. Direct interaction of endothelial nitric-oxide
synthase and caveolin-1 inhibits synthase activity. J Biol Chem 1997; 272:
9.Kashiwagi M, Shinozaki M, Hirakata H et al. Locally activated renin-
angiotensin system associated with TGF-beta1 as a major factor for renal
injury induced by chronic inhibition of nitric oxide synthase in rats. J Am
Soc Nephrol 2000; 11: 616–624.
10.Keynan S, Hirshberg B, Levin-Iaina N et al. Renal nitric oxide production
during the early phase of experimental diabetes mellitus. Kidney Int 2000;
58: 740–747.
11.Kiff RJ, Gardiner SM, Compton AM et al. The effects of endothelin-1 and
NG-nitro-L-arginine methyl ester on regional haemodynamics in
conscious rats with streptozotocin-induced diabetes mellitus. Br J
Pharmacol 1991; 103: 1321–1326.
12.Hohenstein B, Hugo CP, Hausknecht B et al. Analysis of NO-synthase
expression and clinical risk factors in human diabetic nephropathy.
Nephrol Dial Transplant 2008; 23: 1346–1354.
13.Zhao HJ, Wang S, Cheng H et al. Endothelial nitric oxide synthase
deficiency produces accelerated nephropathy in diabetic mice. J Am Soc
Nephrol 2006; 17: 2664–2669.
14.Komers R, Anderson S. Glomerular endothelial NOS (eNOS) expression in
type 2 diabetic patients with nephropathy. Nephrol Dial Transplant 2008;
23: 3037; author reply 3037–3038.
15.Qi Z, Fujita H, Jin J et al. Characterization of susceptibility of inbred mouse
strains to diabetic nephropathy. Diabetes 2005; 54: 2628–2637.
16.Gurley SB, Clare SE, Snow KP et al. Impact of genetic background on
nephropathy in diabetic mice. Am J Physiol Renal Physiol 2006; 290:
17.Goldberg IJ, Dansky HM. Diabetic vascular disease: an experimental
objective. Arterioscler Thromb Vasc Biol 2006; 26: 1693–1701.
18.Bower G, Brown DM, Steffes MW et al. Studies of the glomerular
mesangium and the juxtaglomerular apparatus in the genetically diabetic
mouse. Lab Invest 1980; 43: 333–341.
19.Thum T, Fraccarollo D, Schultheiss M et al. Endothelial nitric oxide
synthase uncoupling impairs endothelial progenitor cell mobilization and
function in diabetes. Diabetes 2007; 56: 666–674.
20.Wu G, Meininger CJ. Nitric oxide and vascular insulin resistance. Biofactors
2009; 35: 21–27.
Jin RC, Loscalzo J. Vascular nitric oxide: formation and function. J Blood
Med 2010; 2010: 147–162.
Baylis C. Nitric oxide deficiency in chronic kidney disease. Am J Physiol
Renal Physiol 2008; 294: F1–F9.
Veelken R, Hilgers KF, Hartner A et al. Nitric oxide synthase isoforms and
glomerular hyperfiltration in early diabetic nephropathy. J Am Soc
Nephrol 2000; 11: 71–79.
Mohan S, Reddick RL, Musi N et al. Diabetic eNOS knockout mice develop
distinct macro- and microvascular complications. Lab Invest 2008; 88:
Kanetsuna Y, Takahashi K, Nagata M et al. Deficiency of endothelial
nitric-oxide synthase confers susceptibility to diabetic nephropathy
in nephropathy-resistant inbred mice. Am J Pathol 2007; 170:
Komers R, Anderson S. Paradoxes of nitric oxide in the diabetic kidney.
Am J Physiol Renal Physiol 2003; 284: F1121–F1137.
Guzik TJ, Mussa S, Gastaldi D et al. Mechanisms of increased vascular
superoxide production in human diabetes mellitus: role of NAD(P)H
oxidase and endothelial nitric oxide synthase. Circulation 2002; 105:
Kobayashi T, Taguchi K, Nemoto S et al. Activation of the PDK-1/Akt/eNOS
pathway involved in aortic endothelial function differs between
hyperinsulinemic and insulin-deficient diabetic rats. Am J Physiol Heart
Circ Physiol 2009; 297: H1767–H1775.
Liu J, Wei S, Tian L et al. Effects of endomorphins on human umbilical vein
endothelial cells under high glucose. Peptides 2011; 32: 86–92.
Kim HW, Lim JH, Kim MY et al. Long-term blockade of vascular
endothelial growth factor receptor-2 aggravates the diabetic renal
dysfunction associated with inactivation of the Akt/eNOS-NO axis.
Nephrol Dial Transplant 2011; 26: 1173–1188.
Ballermann BJ. Glomerular endothelial cell differentiation. Kidney Int 2005;
67: 1668–1671.
Satchell SC, Braet F. Glomerular endothelial cell fenestrations: an integral
component of the glomerular filtration barrier. Am J Physiol Renal Physiol
Thomas SR, Witting PK, Drummond GR. Redox control of endothelial
function and dysfunction: molecular mechanisms and therapeutic
opportunities. Antioxid Redox Signal 2008; 10: 1713–1765.
Li H, Wallerath T, Munzel T et al. Regulation of endothelial-type NO
synthase expression in pathophysiology and in response to drugs.
Nitric Oxide 2002; 7: 149–164.
Searles CD. Transcriptional and posttranscriptional regulation of
endothelial nitric oxide synthase expression. Am J Physiol Cell Physiol
Luo Z, Fujio Y, Kureishi Y et al. Acute modulation of endothelial Akt/PKB
activity alters nitric oxide-dependent vasomotor activity in vivo. J Clin
Invest 2000; 106: 493–499.
Mount PF, Kemp BE, Power DA. Regulation of endothelial and myocardial
NO synthesis by multi-site eNOS phosphorylation. J Mol Cell Cardiol 2007;
42: 271–279.
Komers R, Schutzer WE, Reed JF et al. Altered endothelial nitric oxide
synthase targeting and conformation and caveolin-1 expression in the
diabetic kidney. Diabetes 2006; 55: 1651–1659.
Xu J, Xie Z, Reece R et al. Uncoupling of endothelial nitric oxidase
synthase by hypochlorous acid: role of NAD(P)H oxidase-derived
superoxide and peroxynitrite. Arterioscler Thromb Vasc Biol 2006; 26:
Alp NJ, Channon KM. Regulation of endothelial nitric oxide synthase by
tetrahydrobiopterin in vascular disease. Arterioscler Thromb Vasc Biol
2004; 24: 413–420.
Munzel T, Daiber A, Ullrich V et al. Vascular consequences of endothelial
nitric oxide synthase uncoupling for the activity and expression of the
soluble guanylyl cyclase and the cGMP-dependent protein kinase.
Arterioscler Thromb Vasc Biol 2005; 25: 1551–1557.
Schmidt TS, Alp NJ. Mechanisms for the role of tetrahydrobiopterin in
endothelial function and vascular disease. Clin Sci (Lond) 2007; 113:
Meininger CJ, Marinos RS, Hatakeyama K et al. Impaired nitric oxide
production in coronary endothelial cells of the spontaneously diabetic
BB rat is due to tetrahydrobiopterin deficiency. Biochem J 2000; 349:
Kohli R, Meininger CJ, Haynes TE et al. Dietary L-arginine
supplementation enhances endothelial nitric oxide synthesis in
streptozotocin-induced diabetic rats. J Nutr 2004; 134: 600–608.
Kidney International (2012) 82, 1176–1183
original article
H Cheng et al.: Sepiapterin protects against glomerular injury
Page 8
45.Pannirselvam M, Simon V, Verma S et al. Chronic oral supplementation
with sepiapterin prevents endothelial dysfunction and oxidative stress
in small mesenteric arteries from diabetic (db/db) mice. Br J Pharmacol
2003; 140: 701–706.
Heitzer T, Krohn K, Albers S et al. Tetrahydrobiopterin improves
endothelium-dependent vasodilation by increasing nitric oxide activity in
patients with Type II diabetes mellitus. Diabetologia 2000; 43: 1435–1438.
Ferre J, Naylor EW. Sepiapterin reductase in human amniotic and skin
fibroblasts, chorionic villi, and various blood fractions. Clin Chim Acta
1988; 174: 271–282.
Coronel I, Arellano-Mendoza MG, del Valle-Mondragon L et al.
L-arginine and antioxidant diet supplementation partially restores nitric
oxide-dependent regulation of phenylephrine renal vasoconstriction
in diabetics rats. J Ren Nutr 2010; 20: 158–168.
Bello E, Caramelo C, Martell N et al. Impairment of renal vasodilation with
l-arginine is related to more severe disease in untreated hypertensive
patients. Hypertension 2001; 38: 907–912.
Burton BK, Nowacka M, Hennermann JB et al. Safety of extended
treatment with sapropterin dihydrochloride in patients with
phenylketonuria: results of a phase 3b study. Mol Genet Metab 2011;
103: 315–322.
Puff R, Dames P, Weise M et al. Reduced proliferation and a
high apoptotic frequency of pancreatic beta cells contribute to
genetically-determined diabetes susceptibility of db/db BKS mice.
Horm Metab Res 2011; 43: 306–311.
Leiter EH, Le PH, Coleman DL. Susceptibility to db gene and
streptozotocin-induced diabetes in C57BL mice: control by gender-
associated, MHC-unlinked traits. Immunogenetics 1987; 26: 6–13.
Morrow JD, Roberts 2nd LJ. Mass spectrometric quantification of
F2-isoprostanes in biological fluids and tissues as measure of oxidant
stress. Methods Enzymol 1999; 300: 3–12.
54. Morrow JD, Zackert WE, Yang JP et al. Quantification of the major
urinary metabolite of 15-F2t-isoprostane (8-iso-PGF2alpha) by a stable
isotope dilution mass spectrometric assay. Anal Biochem 1999; 269:
Takemoto M, Asker N, Gerhardt H et al. A new method for large
scale isolation of kidney glomeruli from mice. Am J Pathol 2002;
161: 799–805.
Akis N, Madaio MP. Isolation, culture, and characterization of
endothelial cells from mouse glomeruli. Kidney Int 2004; 65:
Korda M, Kubant R, Patton S et al. Leptin-induced endothelial
dysfunction in obesity. Am J Physiol Heart Circ Physiol 2008; 295:
Cheng HF, Harris RC. Cyclooxygenase-2 expression in cultured cortical
thick ascending limb of henle increases in response to decreased
extracellular ionic content by both transcriptional and post-transcriptional
mechanisms. Role of p38-mediated pathways. J Biol Chem 2002; 277:
Chen S, Apostolova MD, Cherian MG et al. Interaction of endothelin-1
with vasoactive factors in mediating glucose-induced increased
permeability in endothelial cells. Lab Invest 2000; 80: 1311–1321.
Bonner SM, O'Sullivan MA. Endothelial cell monolayers as a model
system to investigate dengue shock syndrome. J Virol Methods 1998;
71: 159–167.
This work is licensed under the Creative Com-
Works 3.0 Unported License. To view a copy of this license,
visit http://creativecommons.org/licenses/by-nc-nd/3.0/
Kidney International (2012) 82, 1176–1183
H Cheng et al.: Sepiapterin protects against glomerular injury
original article


text:
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Special Report: India's World Cup fever (Aired: June 2006)
Oct 7, 2013
I love India, Kerala: Maradona
Oct 24, 2012
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Jun 14, 2010
Viva Kerala revitalises football
Nov 1, 2007
Indian Super League: Kerala Blasters Down FC Pune City 2-1, Register First Win of Tournament
Press Trust of India | Thursday October 30, 2014
Kerala Blasters have overtaken FC Pune City in the Indian Super League standings and are placed fifth in the table led by Atletico de Kolkata.
As It Happened - FC Pune City vs Kerala Blasters FC
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Indo-AsianNews Service | Wednesday October 29, 2014
After a comprehensive victory against Goa, FC Pune City is confident and will look to display another great performance in front of a roaring home crowd.
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Press Trust of India | Sunday October 26, 2014
Atletico de Kolkata maintained their top position in the Indian Super League table with 11 points from five games while Kerala Blasters earned their first point in their third game.
Indian Super League, Live Updates: Table-Toppers Atletico de Kolkata Clash With Kerala Blasters
AshishMaggo | Sunday October 26, 2014
Indian Super League table-toppers Atletico de Kolkata will clash with Kerala Blasters, who are languishing right at the bottom. Can ATK stretch their lead further?
ISL: Top-Placed Atletico de Kolkata Square Off Against Kerala Blasters
Press Trust of India | Saturday October 25, 2014
Atletico de Kolkata are three points clear of NorthEast United FC in the Indian Super League points table.
Indian Super League: Bernard Mendy Magic Wins it for Chennaiyin FC Against Kerala Blasters
Press Trust of India | Tuesday October 21, 2014
Chennaiyin FC now have two wins from as many matches while Kerala Blasters are at the bottom of the table having lost both their games in the inaugural Indian Super League.
Amitabh Bachchan, Rajinikanth Combine Forces, Lead Cheer for Chennaiyin F.C.
NDTVSports | Wednesday October 22, 2014
Between the two, Amitabh Bachchan and Rajnikanth command a billion fans. On the football field though, the mega stars were content being ordinary - and often noisy - supporters themselves.
Indian Super League Highlights: Mendy Scores Stunning Goal as Chennaiyin FC Beat Kerala Blasters 2-1
AshishMaggo | Wednesday October 22, 2014
The South Indian derby between Chennaiyin FC and Kerala Blasters resulted in a 2-1 win for home side Chennai, who found the winner in the second half after the scores were level for a while. The mind-boggling goal from Mendy proved to be the decider ...
Indian Super League: Chennaiyin FC Host Kerala Blasters FC, Eye Second Win
Press Trust of India | Tuesday October 21, 2014
Chennaiyin FC had started their campaign with a win over FC Goa while Kerala Blasters FC went down to Northeast United FC in their tournament opener away for home.
Subroto Cup: Jose Ricardo's Solitary Effort Puts Santo Antonio in Semis
Press Trust of India | Thursday October 16, 2014
Brazilian team Colegio Estadul Santo Antonio will meet Indonesia in the semifinals, to be played on October 18. The winner of that match will then take on finalists MSP School from Kerala on October 20.
Atletico de Kolkata Will be our Toughest Rivals, Says NorthEast United FC Co-Owner John Abraham
Indo-AsianNews Service | Tuesday October 14, 2014
NorthEast United FC are scheduled to host Atletico de Kolkata on October 16 and FC Goa on October 19. They pulled off a hard fought 1-0 victory against Kerala Blasters FC on Monday.
Indian Super League: Spanish Striker Koke Gives NorthEast United 1-0 Win Over Kerala Blasters
Press Trust of India | Monday October 13, 2014
A 45th minute strike from Koke sealed the deal for NorthEast United as they did well to hold on to their slender lead in the action-packed second-half at the Indira Gandhi Stadium in Guwahati.
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Press Trust of India | Monday October 13, 2014
Sachin Tendulkar is one of the co-owners of the ISL franchise Kerala Blasters FC which is being led by former England goalkeeper David James.
MSP Kerala Reach Subroto Cup Final
Press Trust of India | Monday October 13, 2014
Understandably, both the teams played cautiously without taking any goal-bearing initiatives, much to the chagrin of their own benches and the spectators. From the outset it was evident that the two teams would adopt a defence-first approach and look...
As It Happened, ISL Match 2 - NorthEast United FC 1-0 Kerala Blasters FC: Koke's Strike Sees NorthEast United FC Get Off To A Flyer
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PrakashGovindasreenivasan | Monday October 13, 2014
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Sachin Tendulkar is going to share his massive international experience with Kerala Blasters ahead of their ISL opener against NorthEast United FC in Guwahati.
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Press Trust of India | Sunday October 12, 2014
As he walked into the grounds of the Yuva Bharati Krirangan stadium in Kolkata, Sachin Tendulkar got a standing ovation from the 68000 spectators present there.


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He won't win the Heisman, but it looks like USC safety Taylor Mays is going to enjoy his senior year:Powered by Sidelines
Taylor Mays and the USC Song Girls
About HeismanpunditChris Huston, A.K.A. 'The Heisman Pundit', is a Heisman voter and the creator and publisher of Heismanpundit.com, a site dedicated to analysis of the Heisman Trophy and college football. Dubbed "the foremost authority on the Heisman" by Sports Illustrated, HP is regularly quoted or cited during football season in newspapers across the country. He is also a regular contributor on sports talk radio and television.
One Response to Taylor Mays and the USC Song Girls
Anonymous June 9, 2009 at 5:48 pm #
When I see Damian Williams, I think "Darkness! Darkness is spreading!"
The HP Heisman Watch
The latest Heisman Straw Poll
Total points, (with first-place votes in parentheses)
1. Marcus Mariota, QB, Oregon — 23 (7)
2. Todd Gurley, RB, Georgia — 18 (3)
3. Kenny Hill, QB, Texas A&M — 7
About The Author
Chris Huston, A.K.A. 'The Heisman Pundit', is the creator and publisher of Heismanpundit.com, a site dedicated to analysis of the Heisman Trophy and college football.
HP's Plaudits
"The Heisman's Foremost Authority..." -- Sports Illustrated.
"Heisman Pundit knows what he's talking about ...
which is more than I can say of some pundits."--Fox Sports's Stewart Mandel
"Heisman Pundit isn't just the authoritative site for all things Heisman...it's also an engaging, highly-opinionated place for college football discourse."--Fox Sports's Bruce Feldman
"Heisman Pundit is the quintessential source for analysis of the greatest individual trophy in the sporting world."--The Big Ten Network's Tom Dienhart
"Indispensable."--SI's Austin Murphy
"...definitely required reading."--Those nutjobs at EDSBS
- Oh, what a wonderful Heisman race October 2, 2014
- Marcus Mariota leads HeismanPundit's Straw Poll for fourth week in a row September 30, 2014
- Looking ahead to the 2015 Heisman race September 29, 2014
- The HP Heisman Watch, week 5 September 29, 2014
- This week in Heisman history: Vinny Testaverde shines against Oklahoma September 26, 2014
- Kevin: I certainly hope you're wrong, with no disrespect...
- Heismanpundit: 1. Winston is already done due to Heismandment No....
- The only reason the Heisman Trophy matters. | TheDawgSpot.com: […] It's been a long time since I̵...
- Heismanpundit: This list doesn't mean these are the only guys wh...
- Aaron: Two thoughts: 1. I trust that the list also ass...


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T-Mobile is launching the BlackBerry Pearl 8220 Flip today. The Flip will come in two versions. The black version of the 8220 will be appearing online and in T-Mobile stores today. Another dark red version will follow on Monday, October 20, 2008. Both versions will sell for $149.99 with a two-year agreement.
T-Mobile launches BlackBerry Pearl 8220 Flip today
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At around 3 p.m., Anderson County firefighters, EMS and the sheriff's office responded to a large hay barn fire on Hickory Grove Road. At 4:30 p.m., Firefighters were still working to get the fire under control.
So often we hear that fall or winter is a particularly good time to do certain things in the landscape. This is usually due to the fact that plants have entered into winter dormancy.
Well, it's here. I'm truly surprised that there weren't parties all over the county last night. Why? There's a bunch of you that love fall. You love the colors and the temperatures. It's probably my second favorite season, only because it's typically warmer than spring.
It's a myth that turning off the lights isn't necessary. The truth is that it depends on what type of lighting you have and how long the light will be off before you need it again.
This column may bring satisfaction to some who read it because they were right, and irritation to those who were having the all too familiar disagreement about the subject.
To say that I am zealous about fall is an understatement. I love Starbucks Pumpkin Spice lattes and eagerly await its arrival every year, the weather is perfect, the leaves changing colors, apple cider … need I say more? But mostly, fall is my favorite season because it has way of making me feel nostalgic.
By Ben Carlson
Column as I see 'em …
The same organization that oversees the (public) education of your children wrote the following, which would be sort of funny were it not such a serious subject:
"The candidate who wishes to be elected to the school council as a minority representative on the school council must be the minority. However, if a person declares himself/herself a minority, he or she should be considered so for the purpose of fulfilling the requirements of minority council member."
By Shelley Spillman
News staff
Cat cages at the Anderson Humane Society are filled with two and sometimes as many as four cats. Currently, there are 33 cats housed at the humane society, about 40 cats in foster homes and 28 at animal control. The agency was overrun with cats two weeks ago when 30 cats were brought in one day. Being a small facility, the humane society can't take on the influx of cats alone.
In a 4-2 vote, Lawrenceburg City Council approved the second reading of 2014 real and personal property taxes Tuesday of last week.
The city's real estate property taxes were set from $1.90 per $1,000 of assessed valuation to $1.95, which is expected to produce $1,050,339 in revenue. The personal property tax rates were set from $2.29 per $1,000 assessed valuation to $3.07, which is expected to produce $80,985 in revenue.
Furnished by
the Kentucky National Guard
FRANKFORT — When the Kentucky National Guard describes itself as a family organization, it has the proof to back it up. Chief Warrant Officer Jennifer Maggard's story is a prime example.
September 12 will remain a special date for the family as Lawrenceburg resident Maggard was promoted to chief warrant officer three in front of friends and family, just before she swore her younger sister Logan Green into the Kentucky Guard.
Copyright © 2014
The Anderson News is your source for local news, sports, events, and information in Lawrenceburg, KY and the surrounding area.


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It's our Interior Design Magazine!
The hottest interior design ideas, fab finds for the home and the latest in interiors trends. We're always looking for new finds, designers to feature or anything else lovely for the home, so if you've seen something gorgeous and would like it featured in our interior design magazine, please get in touch!
Editors Choice: Tempting tea towels
by Alexandra Thomas on August 17, 2014 | 0 comments
Brighten up your kitchen with our pick of fantastic tea towels.
I'm yet to meet anyone who actively enjoys washing up but, if you're not blessed with a machine that does it for you, then investing in some stylish tea towels will hopefully, at least, make the drying up a little more bearable.
We have some super tea towels here at Furnish, all great at adding colour and pattern to your kitchen.
This Robots Tea Towel by Kukka will charm all ages while this Bear Tea Towel in Magenta by Artstar is equally delightful.
For those who prefer things to be a little more simple, this Red Heart Tea Towel by Red Candy has a classic colour combination of red and white, and will suit both contemporary and classic schemes.
Meanwhile, this Metamorphosis Tea Towel features a striking owl design, which is also available in matching cups, saucers and mugs.
Shop the look...
Aztec Cushions
by Alexandra Thomas on August 14, 2014 | 0 comments
Introduce pattern and colour with these Aztec Cushions, £45.
Aztec prints are big fashion news at the moment so it's no surprise, as with many other catwalk trends, that these bold, vibrant patterns are also making an appearance in the home.
These Aztec Cushions by The French Bedroom Company are ideal for brightening up a plain sofa or giving an oomph to your bed linen, or you could even use them outside on your garden furniture.
Each cushion in this set of three has a slightly different pattern and uses different colour combinations, allowing you to use them together as a bold style statement or singularly with plain cushions in complementary shades.
Shop the look...
Squirrel & Nut Deckchair
by Alexandra Thomas on August 12, 2014 | 0 comments
Sit back and relax with this Squirrel & Nut Deckchair, £135.
There's possibly nothing that screams British summertime more than the classic deckchair.
Perfect for relaxing at home or on the beach, these simple chairs have been around forever, and are ideal for an afternoon snooze in the sun.
We have a great selection of deck chairs here at Furnish but one of my favourites is this Squirrel & Nut Deckchair by Amara Living.
With a hardwood frame and cotton canvas sling, this deckchair is extremely cute with its squirrels and acorns.
All you need now is a glass of Pimms, a gorgeous summer's day and a good book. Perfect!
Shop the look...
Colour watch: Citrus shades
by Alexandra Thomas on August 10, 2014 | 0 comments
Add some zing to your home with our selection of brightly coloured buys.
As we approach the middle of summer, whether in rain or in shine, there couldn't be a better time to inject some fresh, zingy, citrus colours to your scheme.
For lovers of lime, this Bugatti Citrus Juicer by Red Candy will instantly brighten up any kitchen worktop, as will this High And Dry Dishrack by Design My World.
Orange obsessives are catered for too; this retro-inspired Orbit Side Table by Naken Interiors is perfect for any room in the home, while this Larry Orange Outdoor Bean Bag by Neofurn is ideal for the garden.
Yellow is possibly the ultimate summer colour, so if you want to add some sunshine brightness, this Kandinsky Cuckoo Clock by Red Candy will become the focal point of any room, while this Eden Yellow Industrial Cage Pendant Light by Alexander and Pearl adds colour to a more industrial scheme.
Shop the look...
Vogue Copper & Grey Lounge Chair
by Alexandra Thomas on August 10, 2014 | 0 comments
Enjoy classic vintage style with this Vogue Copper & Grey Lounge Chair, £1,395.
Regular readers will know that I am completely obsessed with chairs. I have as many as I can possibly squeeze into my home, I have prints of classic chairs on the wall and I even have a whole shelf on my bookcase dedicated to the design of chairs.
So, while I don't have the room for a new chair, that doesn't mean I can't look, and my new favourite is this Vogue Copper & Grey Lounge Chair by Alexander and Pearl.
With a striking, on-trend copper base and simple yet comfortable grey upholstery, this statement piece will look fantastic in the bedroom or living space, particularly when teamed with a bright neon pink or yellow cushion.
Also available as an armchair model.
Shop the look...
Laundry Punch Bag
by Alexandra Thomas on August 09, 2014 | 0 comments
Keep dirty laundry stashed away with this Laundry Punch Bag, £20.
Laundry bins are the bugbear of my life because, although they're completely necessary, they're not always the most attractive of items. So much so, I hide mine away in a cupboard so it never sees the light of day.
Which is why I was delighted to come across this fab Laundry Punch Bag by Rose & Grey.
Designed to look like a punch bag, and working as one when full, it is great for easing the frustrations of laundry day.
Also ideal as a gift for older children or teenagers to encourage them to keep their dirty laundry tidied away.
Shop the look...
Geometric Beech Suspension Light
by Alexandra Thomas on August 05, 2014 | 0 comments
Update your lighting with a Geometric Beech Suspension Light, £39.90.
While it is still summer, the days are steadily getting shorter, so there's no better time to sort out your lighting scheme in time for autumn.
Lighting is a notoriously tricky element of interior design to get right, which is why many get it wrong.
A simple trick is to make sure you have more than one lighting source in a room (e.g. table lights, pendant lamps, etc.) so you have different lights for various purposes. Adding a dimmer switch is a great idea too if you want to create varying atmospheres.
This Geometric Beech Suspension Light by Cachette has just jumped to the top of my wish list and I can't wait to get it in my home.
Available in a range of shapes, these lights have been crafted from solid wood and will look equally stylish used alone or grouped together.
Shop the look...
Kenwood kMix Brights Stand Mixer
by Alexandra Thomas on August 01, 2014 | 0 comments
Add some retro style to your kitchen with this Kenwood kMix Brights Stand Mixer, £350.
I'll be honest – I don't really need a food mixer. I definitely don't bake enough cakes or make enough bread to warrant one, and I already have a whole host of "must-have" kitchen gadgets I never use.
But I could be persuaded to go all Mary Berry if I had this gorgeous Kenwood kMix Brights Stand Mixer by Red Candy.
Deliciously retro, it would be a welcome addition to my kitchen for its looks alone, but if you are a whizz when it comes to meringues, then it's good to know it's got loads of functions to make whipping up tasty treats an absolute breeze.
Also available in raspberry red, orange, pink and yellow.
Shop the look...
Brown Open Weave Bench
by Alexandra Thomas on July 31, 2014 | 0 comments
Relax in style with this Brown Open Weave Bench, £197.
I've always been a bit 'meh' when it comes to having benches in the garden; for me they can either look like they belong in a park or are too ornate to fit in with the sleek style I favour.
However, from our fab range of garden benches here at Furnish, I've found (and now have my heart set on) this Brown Open Weave Bench by Lover's lounge.
With a contemporary, yet intricately woven back, this bench has sleek legs and a generous cushion allowing you to comfortably relax in your outdoor space and enjoy the sun.
Shop the look...
Trend Watch: Super stripes
by Alexandra Thomas on July 27, 2014 | 0 comments
Give your home a smart look with our round up of striped goodies.
Stripes are a perennial favourite, however, they always make me think of summer. Maybe it's all those Breton tops and denim shorts combos you see when the sun decides to shine, or perhaps it's because they're particularly nautical in style.
I adore a stripe in the home as well as in fashion; they're easy to incorporate into any room from kitchen to bathroom, and suit any scheme whether contemporary or classic.
Great for keeping your bits and pieces hidden and being super stylish at the same time, I love this Paulie Orange & White Stripe Storage Cube Trunk by Alexander and Pearl, while this Black Stripe Cushion by Naken Interiors will instantly update any bed, chair or sofa.
This Swish Black and White Striped Armchair by Lover's lounge offers a subtle way to incorporate the trend and can easily be combined with other patterns without appearing too garish, while this Multi-Striped Chest of Drawers by Out There Interiors is ideal for those who are bold when it comes to decorating the home.
Shop the look...
Joseph Joseph Baking Gift Set
by Alexandra Thomas on July 27, 2014 | 0 comments
Bake beautifully with this Joseph Joseph Baking Gift Set, £35.
If you love baking and could easily be a contestant on the next series of The Great British Bakeoff, chances are you have all the paraphernalia you need.
However if, like me, you're just starting out then you need to make sure you have the right equipment.
We have a whole host of baking and kitchenware here on Furnish from mixing bowls to pretty storage tins but I've got my heart set on this super cool Joseph Joseph Baking Gift Set by Red Candy.
With an adjustable rolling pin, clever spatula and pastry brush, and all-important timer, this intelligent kit will ensure Mary Berry et al will be desperate to taste your wares.
Shop the look...
Scion Zig Zag Soap Dispenser
by Alexandra Thomas on July 26, 2014 | 0 comments
Add some pattern and colour with this Scion Zig Zag Soap Dispenser, £13.
I'm always looking for ways to brighten up my fairly dull bathroom, whether by adding colourful patterned towels or using wall stickers to jazz up the walls.
If you're looking for a quick and easy solution, then this Scion Zig Zag Soap Dispenser by Amara Living is ideal.
Bright, bold and definitely beautiful, it will add colour and pattern to your basin in a flash, and mean you don't have to worry about unsightly hand wash dispensers.
Also available in the range is a bath mat, tumbler, soap dish and toilet brush which look great teamed with a chic white bathroom.
Shop the look...
Quick Shop: Keeping Cool
by Alexandra Thomas on July 19, 2014 | 0 comments
Keep your cool in the summer months with these inspired buys.
No sooner is a heatwave announced then we all start calling for rain. Turns out when it comes to weather; you can't please all of the people all of the time.
So, if the heat is getting to you, we have some great products that will ensure you keep your cool when everyone around you is losing theirs.
If picnics and al fresco dining are your thing then you'll need this Menu Picnic Wine Cooler by Gifts with Style, which will keep your bottle of plonk nicely chilled. Meanwhile, the TEA-JAY Ice Tea Maker Kit by Naken Interiors is a unique item perfect for iced tea and other chilled mixed drinks.
Naturally, you'll want some ice cream or sorbet in this weather so serve it in style with this funky Alessi Big Love Ice Cream Bowl by Red Candy, which is available in a myriad of bright colours.
And finally, for a bit of fun, we couldn't resist this Newgate Ice Lolly Wall Clock, also by Red Candy, which is perfect for adding some summer charm to your home.
Shop the look...
Anorak Melamine Rolling Hills Plate
by Alexandra Thomas on July 19, 2014 | 0 comments
Enjoy al fresco dining with this Anorak Melamine Rolling Hills Plate, £5.50.
The forecast is looking good and with the sun comes the chance to get outside for some al fresco dining, either in your garden or with a picnic.
Unless you're really posh, you probably won't want to risk your best china, which is why melamine makes an ideal alternative.
We have a great range of melamine goodies here at Furnish, with this Anorak Rolling Hills Melamine Plate by Amara Living really catching my eye.
Simple yet ever so sweet, these durable plates are ideal for everything from salads to sandwiches.
And if you're looking for bowls and beakers too you'll be pleased to know they are also available in this range.
Shop the look...
Brick Vase
by Alexandra Thomas on July 16, 2014 | 0 comments
Create a simple floral display with this Brick Vase, £30.
As a lover of fresh flowers in the home, I have amassed a rather impressive collection of vases, however, after recently deciding that mine were all rather girly, I have fallen for this Brick Vase by Naken Interiors.
Sturdy and stylish, it will definitely add a masculine look to my floral displays, and I adore the juxtaposition of something so solid containing something so delicate.
Great for teaming with glass designs to create an eclectic centrepiece for a dining table, this vase will look equally good alone on a sideboard.
Shop the look...
Editors Choice: Bar Stools
by Alexandra Thomas on July 13, 2014 | 0 comments
Make the most of your kitchen space with our great selection of bar stools.
A bar stool is an great piece of kit and can instantly transform otherwise dead space in your kitchen.
And there's no need for an expansive breakfast bar as, with just the end of a worktop, you have the perfect place to perch whether it's for breakfast or an after-work cocktail.
With an array of bar stools to choose from here at Furnish, there's something to suit every style and budget.
Perfect for adding a touch of girly chic to any kitchen/diner space, this Luke Acrylic Pink Bar Stool by Neofurn will instantly add a pop of colour, while this Manhattan Wood & Iron Bar Stool by Alexander & Pearl is ideal for adding a subtle industrial touch.
For those who prefer to keep things sleek and simple, this Chrome Bar Stool by London Cows is sure to be a hit, while design lovers will adore this Raft Bar Stool by Naken Interiors.
Shop the look...
Rustic Grey Heart Mirror
by Alexandra Thomas on July 12, 2014 | 0 comments
Add some romance to your home with this Rustic Grey Heart Mirror, £40.
Mirrors offer a great way to instantly lift a room and rejuvenate a plain wall. They also help increase the feeling of light and space, which is particularly useful if you want to brighten up a gloomy corner or area.
We have a great range of mirrors to choose from here at Furnish from elaborate over-mantel designs to sleek, full length options, which are perfect for a contemporary interior.
I love this Rustic Grey Heart Mirror by Primrose & Plum that has been crafted from metal and finished with a rope hanging strap.
Simple and understated, yet ever-so stylish, this mirror will work well in every room in the home from bathroom to hallway.
Shop the look...
Vintage Martini Cocktail Glasses
by Alexandra Thomas on July 08, 2014 | 0 comments
Enjoy stylish cocktails with these Vintage Martini Cocktail Glasses, £17.88.
While I love vintage furniture, I have to admit to feeling a little bit squeamish about eating and drinking from vintage crockery and glasses.
So, in my search for some new, vintage-style tableware I am thrilled to have come across these gorgeous Vintage Martini Cocktail Glasses by Solavia, which are just what I've been looking for.
Of course they're perfect for serving all different kind of martinis, but they're also ideal for a Cosmopolitan or (my personal favourite) a Negroni.
Best of all they're much more affordable than they look, which means they won't break the bank.
Shop the look...
Orange Comic Boom Cushion
by Alexandra Thomas on July 07, 2014 | 0 comments
Add some super cool chic with this Orange Comic Boom Cushion, from £34.
While we take our interiors very seriously here at Furnish, we like to have fun too, which is why this Orange Comic Boom Cushion by Red Candy is one of my favourite recently-added products.
Taking inspiration from comic books, this cushion is one of a collection of ten pop art-style cushions, which are all perfect for adding a brilliant hit of colour to your home. Try using just the one or add a selection of your favourites for maximum impact.
Great for jazzing up a neutral sofa, these cushions are also ideal for a little one's room.
Shop the look...
Parlane Beach Hut Hook
by Alexandra Thomas on July 03, 2014 | 0 comments
Add some sea-side style with this Parlane Beach Hut Hook, £8.
Some people love climbing hills, some love skiing, but my passion is being at the beach.
I'm convinced it has something to do with living in Cornwall for three years or perhaps, as a Pisces, I just love to be by the sea. But, no matter whether it's tropical or wind-ravaged in winter, I'm happiest when I'm at the coast.
Which means I've got a bit of a thing for coastal style and nautical touches, and that is why I love this Parlane Beach Hut Hook by John Lewis.
Irresistibly sweet without being too cute, it's perfect for adding a nautical touch to your kitchen or bathroom.
Shop the look...


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It's Not Gas
... to update, a truck inspection to do, WD-40 to buy, oil to check.... Pet peeve: restrooms with no paper towels Why? Well..... #1: the blowers don't dry your hands #2: you (and everyone else) have to touch everything on the way out #3: if you need to wash your face you have to dry it on your dress #4: the blowers are loud One more thing. I can't figure out why automatic flush toilets always flush while I'm still sitting on ...
Home Sweet Home
... Counting all modes of transportation (cars, big and little planes, train, motor home, buses, and ship---see pics) we covered a total of 10,295 miles. Our legs feel like you could double that if walking is thrown into the mix. Impressive for us but still relatively meager when compared to our son Brian who reminded us that during our 43 day adventure he traveled through 14 airports from SC to Italy to Netherlands to SC to Florida to SC to ...
Blue Ridge Mts and East to SC and Back to GA
... the type of place that you order and pay up front. When we walked up to the cash register, we were greeted by a guy with a friendly face who said, "Ya'll aren't from around
here are you?" (hence the Aggie shirts) We told him our story and we hit it off with him immediately. It turns out he used to play football at USC and is now the owner of the Palmetto Pig. ...
Trip Week--Finally
Ben and I, along with our Tennessee friends, Henry and Tricia McKinnon, have been preparing for this adventure to Alaska for a year and the time is finally here. We fly out to Minneapolis on Wednesday to spend a few days with cousins Cindy and Earl Grothman and then on to Forrest City, Iowa and the Winnebago factory on Saturday. We will begin our adventure on ...
Move in day.
... lonely. We went to walmart with our new friend Lucy who studies here. I already had my sheets so I bought a matress topper because the bed is some weird plastic thing you'd expect to find in a caravan park. I bought some food, a washing basket and some detergent.
After that Lucy dropped us back to our dorms and I begun washing all my clothes from New York. I have to pay $1.25 in quarters for the washer and a $1.25 in quarters for the ...


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text: HR SCREENER JOB DESCRIPTIONS Washroom Attendant Job Description ...Serves patrons of lavatories in store, public building, hotel, or similar establishment by p... FULL JOB DESCRIPTION Printed Circuit Board Assembly Touch up Screener Job Description ...Inspects printed circuit board Pcb assemblies for defects, l... FULL JOB DESCRIPTION Shredder Picker Job Description ...Feeds raw fiber stock into machines that loosen, shred, separate, clean, form nubs, or straight... FULL JOB DESCRIPTION HR Analyst Job Description ...Interact with hr generalists, managers, employees other human resources colleagues to help support various programs for the human resource team. Provide day to day hr support to the business, answering employee and manager questions about hr programs, policies, benefits other hr related items. Assist with hr transactions including benefit changes, processi... FULL JOB DESCRIPTION HR Compensation Benefits And Job Analysis Specialist Job Description ...Ensure company compliance with federal state laws, including reporting requirements. Plan, develop, evaluate, improve, and communicate methods techniques for selecting, promoting, compensating, evaluating, and training workers. Provide advice on the resolution of classification salary complaints. Prepare occupational classifications, job descriptions salary scales. Assist in preparing and maintaining personnel records handbooks. Prepare reports, like organization and flow charts, and career path reports, to summar... FULL JOB DESCRIPTION Cloth shrinking machine Operator Job Description ...Tends machine that shrinks woven or knitted cloth to predetermined size. Mounts roll of cloth on machine, using chain hoist. Threads end of cloth through tension bars, guide feed rollers and through shrinking elements. Observes flow of cloth through machine onto takeup roll, or guides cloth onto shrinking elements to prevent wrinkles uneven winding, or sets speed of elements according to shrinkage-control guide for various cloth styles. Turns valves or moves switc... FULL JOB DESCRIPTION Three dimensional map Modeler Job Description ...Makes plaster relief-map models from which molds are prepared to produce duplicate scale models of land forms. Mixes plaster, water and retarder in container. Punches holes in maps surface, using stippler or scorer so that plaster will adhere. Pours plaster over sea-level contour map, and spreads it evenly over surface with brush or spatula. Covers drainage, river, and city lines with transparent media or leaves border without plaster to preserve lines. Builds mound or ... FULL JOB DESCRIPTION Touch up Screener Printed Circuit Board Assembly Job Description ...Inspects printed circuit board Pcb assemblies for defects, like missing or damaged components, loose connections, or defective solder. Examines Pcbs under magnification lamp compares boards to sample board to detect defects. Labels defects requiring extensive repairs, like missing or misaligned parts, damaged components and loose connections and routes boards to repairer. Performs minor repairs, like cleani... FULL JOB DESCRIPTION Shrinking machine Operator Job Description ...Tends machine that kneads fur felt hat cones to interlock fibers shrink cones by either of following methods. 1 Positions cones on belt that moves cones beneath water sprinkler pipes, between rollers and onto second belt that returns cones to worker. Measures cones, using ruler. Repeats process until cone has shrunk to size s... FULL JOB DESCRIPTION HR Officer Job Description ...Assist the HR Manager in personnel administration, implementation of organization HR policies their follow-up, staffing issues or industrial relations. Propose action plans participate in their deployment. Collect information & responsible for preparing reports in line with organi... FULL JOB DESCRIPTION HR Generalist Job Description ...Resume Duties: Drives talent acquisition initiatives for non-exempt positions. Utilizes applicant tracking system, online job boards, cold-calling and networking to recruit attract talent to the organization. Manages coordinates hourly performance review process. Partner with managers to make sure compensation performance plans are aligned. Manages coordinates hourly employee succession planning including gap analysis, career development planning, and performance differentiation. Administers and manages all company benefits leave programs including medical, dental, 401k, pension, fm... FULL JOB DESCRIPTION Transportation Security Screener Job Description ...Resume Duties: Ask passengers to remove shoes divest themselves of metal objects prior to walking through metal detectors. Close entry areas following security breaches or reopen areas after receiving notification that the airport is secure. Confiscate dangerous items and hazardous materials found in opened bags turn them over to airlines for disposal. Contact leads or supervisors to discuss objects of concern that are not on prohibited object lists. Contact police directly in cases of urgent security issues, using phones or two-way radios. Decide whether baggage that t... FULL JOB DESCRIPTION Pipe thread Inspector Job Description ...Resume Duties: Inspects threads on joints of steel casings, tubings, drillpipes, and other tubular oil well equipment to detect defects and to define conformance to specified tolerances in taper, lead, and threads per inch, using precision gauges instruments. Measures machined threads and outside diameter of pipes to define thread size, taper, th... FULL JOB DESCRIPTION Supervisor Kashruth Job Description ...Resume Duties: Supervises workers engaged in storing, preparing, and cooking meats, poultry, and other foods in restaurants, catering halls, hospitals, or other establishments to make sure observance of Hebrew dietary laws customs. Examines incoming purchases of meat and fowl for making sure that slaughtering selection of meat cuts have been performed according to dietary law. Inspects equipment, utensils and food su... FULL JOB DESCRIPTION Screener Operator Job Description ...Resume Duties: Tends machine that segregates particles of dry materials, like sand, powdered minerals, feed, resins, and chemicals, into batches of uniform size by sifting through one or more oscillating screens. Bolts or clamps screen of specified mesh to frame, using handtools. Pours or shovels specified quantities of material into hopper... FULL JOB DESCRIPTION


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I have the following table in a form that's embedded in a formtastic form (semantic_form_for). Everything I ask to be generated by ruby shows up, but the table gets badly mangled (essentially, the tags NEVER get formed.
The table headers get drawn correctly
There are 14 available_date objects that get passed, and they alternate between having a time value of 1 or 2, so this is just terribly boggling, but probably simple to fix...

<% f.fields_for :available_dates do |ad| %>
<% if ad.object.time == 1 #if this is an early shift, then start the new row %>

<%= ad.object.date.strftime('%a, %b %d, %Y') %>
<%= ad.collection_select(:availability , LookupAvailability.all.collect, :id, :name) %>
<% else #otherwise end the row with just a box%>
<%= ad.collection_select(:availability , LookupAvailability.all.collect, :id, :name) %>

<% end %>
<% end %>
So like I said, the form functions properly, and the objects all get updated and displayed correctly and all that, it's just that the HTML isn't getting echo'd out properly so my table is all mangled. Help!


text:
-since Dec 17, 2011
can you train your hair to be straight?
like how u can train ur hair to curl sumwhat? like i was havin a convo wit a coworker who has natural hair and blow dries it str8 [i mean bone str8 too] every week cuz she doesnt want to deal wit the natural curlish state of it. like isnt that just heat damage and not training it to be str8. and mind you she also wants healthy hair; there is nothin healthy at all bout that. let me kno
- ravies answered: that is def heat damage… no way that be heathly
- susurrus answered: I went thru a brief straight craze and my once-unruly curls have flattened out now, not dry, just dead. is there a way to get my oomph back?!
- xccentric answered: if the right products are used then its not damage
- everybodyluvskay answered: Yes. I had healthy "trained" hair when I was in high school. If you straighten a lot, with low heat and heat protectant, it's not that bad.,
- morotsbiff answered: Hahah, there is no such possibility, my sisters got big curls and i've got nappy hair, i guess if you braid it alot then it somehow…nah
- fivefootnadaa answered: if you do it enough it can flatten your curl pattern…my cousin accidentally did it to her hair. she over flat ironed it too much
- ethiopienne answered: Google "heat training."
- kgchrissy answered: No I don't think you can train it to be str8. My sister is natural and keeps it str8 but when she washes it you can see the heat damage.
- hellzheart answered: To me whenever someone talks about training their hair it just means heat damaging it into submission.
- itisblackandblack answered: they are essentially the same thing. by damaging the hair, one "trains" it to be straight. Training is just a nicer word for damaging.
- hi-imcurrentlyobsessed answered: umm yea heat damage for sure
- liquefieddopeness answered: Noo it's a technique you use. You can blowdry your hair every week and be fine if you use a heat protectant
- sarahsosincere answered: LOL! Yeah, I'm pretty sure that's just heat damage. She's permanently killing her curls.
- kinkycurls posted this
Embracing Natural Kinks && Curls Everywhere! A blog on natural hair, the products that help us deal with our kinks and curls, and the ups and downs of it all.