bluuebunny/crossref_metadata_2025 huggingface dataset:
Fur-Dependent Detoxification of Organic Acids by
rpoS
Mutants during Prolonged Incubation under Aerobic, Phosphate Starvation Conditions
Authors: Guillemet Mélanie L. , Moreau Patrice L.
Published: June 2008

The activity of amino acid-dependent acid resistance systems allows
Escherichia coli
to survive during prolonged incubation under phosphate (P
i
) starvation conditions. We show in this work that
rpoS
-null mutants incubated in the absence of any amino acid survived during prolonged incubation under aerobic, P
i
starvation conditions. Whereas
rpoS
+
cells incubated with glutamate excreted high levels of acetate,
rpoS
mutants grew on acetic acid. The characteristic metabolism of
rpoS
mutants required the activity of Fur (ferric uptake regulator) in order to decrease the synthesis of the small RNA RyhB that might otherwise inhibit the synthesis of iron-rich proteins. We propose that RpoS (σ
S
) and the small RNA RyhB contribute to decrease the synthesis of iron-rich proteins required for the activity of the tricarboxylic acid (TCA) cycle, which redirects the metabolic flux toward the production of acetic acid at the onset of stationary phase in
rpoS
+
cells. In contrast, Fur activity, which represses
ryhB
, and the lack of RpoS activity allow a substantial activity of the TCA cycle to continue in stationary phase in
rpoS
mutants, which decreases the production of acetic acid and, eventually, allows growth on acetic acid and P
i
excreted into the medium. These data may help explain the fact that a high frequency of
E. coli rpoS
mutants is found in nature.
Identification and Utility of FdmR1 as a
Streptomyces
Antibiotic Regulatory Protein Activator for Fredericamycin Production in
Streptomyces griseus
ATCC 49344 and Heterologous Hosts
Authors: Chen Yihua , Wendt-Pienkowski Evelyn , Shen Ben
Published: June 2008

The fredericamycin (FDM) A biosynthetic gene cluster, cloned previously from
Streptomyces griseus
ATCC 49344, contains three putative regulatory genes,
fdmR
,
fdmR1
, and
fdmR2
. Their deduced gene products show high similarity to members of the
Streptomyces
antibiotic regulatory protein (SARP) family (FdmR1) or to MarR-like regulators (FdmR and FdmR2). Here we provide experimental data supporting FdmR1 as a SARP-type activator. Inactivation of
fdmR1
abolished FDM biosynthesis, and FDM production could be restored to the
fdmR1
::
aac(3)IV
mutant by expressing
fdmR1
in
trans
. Reverse transcription-PCR transcriptional analyses revealed that up to 26 of the 28 genes within the
fdm
gene cluster, with the exception of
fdmR
and
fdmT2
, were under the positive control of FdmR1, directly or indirectly. Overexpression of
fdmR1
in
S. griseus
improved the FDM titer 5.6-fold (to about 1.36 g/liter) relative to that of wild-type
S. griseus
. Cloning of the complete
fdm
cluster into an integrative plasmid and subsequent expression in heterologous hosts revealed that considerable amounts of FDMs could be produced in
Streptomyces albus
but not in
Streptomyces lividans
. However, the
S. lividans
host could be engineered to produce FDMs via constitutive expression of
fdmR1
; FDM production in
S. lividans
could be enhanced further by overexpressing
fdmC
, encoding a putative ketoreductase, concomitantly with
fdmR1
. Taken together, these studies demonstrate the viability of engineering FDM biosynthesis and improving FDM titers in both the native producer
S. griseus
and heterologous hosts, such as
S. albus
and
S. lividans
. The approach taken capitalizes on FdmR1, a key activator of the FDM biosynthetic machinery.
Induction by Cationic Antimicrobial Peptides and Involvement in Intrinsic Polymyxin and Antimicrobial Peptide Resistance, Biofilm Formation, and Swarming Motility of PsrA in
Pseudomonas aeruginosa
Authors: Gooderham W. James , Bains Manjeet , McPhee Joseph B. , Wiegand Irith , Hancock Robert E. W.
Published: June 2008

Pseudomonas aeruginosa
is an important opportunistic pathogen that causes infections that can be extremely difficult to treat due to its high intrinsic antibiotic resistance and broad repertoire of virulence factors, both of which are highly regulated. It is demonstrated here that the
psrA
gene, encoding a transcriptional regulator, was upregulated in response to subinhibitory concentrations of cationic antimicrobial peptides. Compared to the wild type and the complemented mutant, a
P. aeruginosa
PAO1
psrA
::Tn
5
mutant displayed intrinsic supersusceptibility to polymyxin B, a last-resort antimicrobial used against multidrug-resistant infections, and the bovine neutrophil antimicrobial peptide indolicidin; this supersusceptibility phenotype correlated with increased outer membrane permeabilization by these agents. The
psrA
mutant was also defective in simple biofilm formation, rapid attachment, and swarming motility, all of which could be complemented by the cloned
psrA
gene. The role of PsrA in global gene regulation was studied by comparing the
psrA
mutant to the wild type by microarray analysis, demonstrating that 178 genes were up- or downregulated ≥2-fold (
P
≤ 0.05). Dysregulated genes included those encoding certain known PsrA targets, those encoding the type III secretion apparatus and effectors, adhesion and motility genes, and a variety of metabolic, energy metabolism, and outer membrane permeability genes. This suggests that PsrA might be a key regulator of antimicrobial peptide resistance and virulence.
Localization of the Germination Protein GerD to the Inner Membrane in
Bacillus subtilis
Spores
Authors: Pelczar Patricia L. , Setlow Peter
Published: June 2008

GerD of
Bacillus subtilis
is a protein essential for normal spore germination with either
l
-alanine or a mixture of
l
-asparagine,
d
-glucose,
d
-fructose, and potassium ions. GerD's amino acid sequence suggests that it may be a lipoprotein, indicating a likely location in a membrane. Location in the spore's outer membrane seems unlikely, since removal of this membrane does not result in a
gerD
spore germination phenotype, suggesting that GerD is likely in the spore's inner membrane. In order to localize GerD within spores, FLAG-tagged GerD constructs were made, found to be functional in spore germination, and detected in immunoblots of spore extracts as not only monomers but also dimers and trimers. Upon fractionation of spore extracts, GerD-FLAG was found in the inner membrane fraction from dormant spores and was present at ∼2,000 molecules/spore. GerD-FLAG in the inner membrane fraction was solubilized by Triton X-100, suggesting that GerD is a lipoprotein, and the protein was also solubilized by 0.5 M NaCl. GerD-FLAG was not processed proteolytically in a
B. subtilis
strain lacking
gerF
(
lgt
), which encodes prelipoprotein diacylglycerol transferase (Lgt), indicating that when GerD does not have a diacylglycerol moiety, signal sequence processing does not occur. However, unprocessed GerD-FLAG still gave bands corresponding to monomers and dimers of slightly higher molecular weight than that of GerD-FLAG from a strain with Lgt, further suggesting that GerD is a lipoprotein. Upon spore germination, much GerD became soluble and then appeared to be degraded as the germinated spores outgrew and initiated vegetative growth. All of these results suggest that GerD is a lipoprotein associated with the dormant spore's inner membrane that may be released in some fashion from this membrane upon spore germination.
Insights into Plant Cell Wall Degradation from the Genome Sequence of the Soil Bacterium
Cellvibrio japonicus
Authors: DeBoy Robert T. , Mongodin Emmanuel F. , Fouts Derrick E. , Tailford Louise E. , Khouri Hoda , Emerson Joanne B. , Mohamoud Yasmin , Watkins Kisha , Henrissat Bernard , Gilbert Harry J. , Nelson Karen E.
Published: June 2008

The plant cell wall, which consists of a highly complex array of interconnecting polysaccharides, is the most abundant source of organic carbon in the biosphere. Microorganisms that degrade the plant cell wall synthesize an extensive portfolio of hydrolytic enzymes that display highly complex molecular architectures. To unravel the intricate repertoire of plant cell wall-degrading enzymes synthesized by the saprophytic soil bacterium
Cellvibrio japonicus
, we sequenced and analyzed its genome, which predicts that the bacterium contains the complete repertoire of enzymes required to degrade plant cell wall and storage polysaccharides. Approximately one-third of these putative proteins (57) are predicted to contain carbohydrate binding modules derived from 13 of the 49 known families. Sequence analysis reveals approximately 130 predicted glycoside hydrolases that target the major structural and storage plant polysaccharides. In common with that of the colonic prokaryote
Bacteroides thetaiotaomicron
, the genome of
C. japonicus
is predicted to encode a large number of GH43 enzymes, suggesting that the extensive arabinose decorations appended to pectins and xylans may represent a major nutrient source, not just for intestinal bacteria but also for microorganisms that occupy terrestrial ecosystems. The results presented here predict that
C. japonicus
possesses an extensive range of glycoside hydrolases, lyases, and esterases. Most importantly, the genome of
C. japonicus
is remarkably similar to that of the gram-negative marine bacterium,
Saccharophagus degradans
2-40
T
. Approximately 50% of the predicted
C. japonicus
plant-degradative apparatus appears to be shared with
S. degradans
, consistent with the utilization of plant-derived complex carbohydrates as a major substrate by both organisms.
Demystified molecular pathology of NUT midline carcinomas
Authors: French Christopher A
Published: June 2008

NUT midline carcinoma (NMC) is a rare, highly lethal cancer that occurs in children and adults of all ages. NMCs uniformly present in the midline, most commonly in the head, neck or mediastinum, as poorly differentiated carcinomas with variable degrees of squamous differentiation. This tumour is defined by rearrangement of the nuclear protein in testis (NUT) gene on chromosome 15q14. In most cases,NUTis involved in a balanced translocation with theBRD4gene on chromosome 19p13.1, an event that creates aBRD4–NUTfusion gene. Variant rearrangements, some involving theBRD3gene, occur in the remaining cases. NMC is diagnosed by detection ofNUTrearrangement by fluorescence in situ hybridisation or reverse transcriptase PCR. Due its rarity and lack of characteristic histological features, most cases of NMC currently go unrecognised.
Human papillomaviruses do not play an aetiological role in Müllerian adenosarcomas of the uterine cervix
Authors: Mumba E , Ali H , Turton D , Cooper K , Grayson W
Published: June 2008

Aim:

To determine if human papillomaviruses (HPVs) play a role in the histogenesis of adenosarcomas of the uterine cervix.

Methods:

Nine archival cases of primary cervical adenosarcoma were studied. The HPV status of the nine histologically proven tumours was investigated by non-isotopic in situ hybridisation (NISH) and PCR. NISH was performed using digoxigenin labelled probes to HPV types 6, 11, 16, 18, 31 and 33. PCR used GP5+/GP6+ primers to the HPV L1 gene.

Results:

Neither the benign epithelial components nor the malignant stromal components of the 9 neoplasms harboured nuclear NISH signals for the HPV types investigated. Amplimers of the HPV L1 gene were not detected by PCR in any of the tumours studied.

Conclusion:

HPVs do not appear to play an aetiological role in cervical adenosarcomas. This suggests that a different histogenetic pathway for this rare tumour type must exist.
Pathological evidence of rhabdomyolysis-induced acute tubulointerstitial nephritis accompanyingLegionella pneumophilapneumonia: Figure 1
Authors: Shimura C , Saraya T , Wada H , Takata S , Mikura S , Yasutake T , Kato J , Kato A , Yamamoto M , Watanabe M , Yokoyama T , Kurai D , Ishii H , Aoshima M , Yamada A , Goto H
Published: June 2008

A case ofLegionella pneumophilapneumonia with rhabdomyolysis-induced acute tubulointerstitial nephritis (ATIN) and prolonged renal dysfunction is presented. The patient was a 54-year-old man, admitted with high-grade fever, ataxia and muscle dysfunction; chest roentgenogram showed multilobular infiltrations.L pneumophilawas detected in his sputum and urine, by PCR and by culture, andL pneumophilapneumonia was diagnosed. Despite antimicrobial treatment, he developed renal failure and rhabdomyolysis. Renal biopsy showed the presence of myoglobin casts that occluded the distal tubuli and tubulointerstitial nephritis, leading to the diagnosis of rhabdomyolysis-induced ATIN. Renal function subsequently normalised, and he was discharged. This is believed to be the first pathological evidence of involvement of rhabdomyolysis in legionellosis-associated ATIN.
Correlation of cyclin D1 transcript levels, transcript type and protein expression with proliferation and histology among mantle cell lymphoma
Authors: Shakir R , Ngo N , Naresh K N
Published: June 2008

Background:

Cyclin D1 expression is central to mantle cell lymphoma (MCL) biology. The cyclin D1 gene produces two forms of mRNA: long (D1L) and short (D1S) versions.

Aims:

To study the relationship between histology, cyclin D1 mRNA (transcript) levels, cyclin D1 transcript type, cyclin D1 protein expression by immunohistochemistry (IHC), and proliferation (Ki-67%).

Methods:

17 MCLs were initially studied for: levels of expression of cyclin D1 transcripts and for cyclin D1 transcript type by reverse-transcriptase PCR; intensity and percentage cyclin D1 protein expression by IHC; and Ki-67% by IHC. The relationship between cyclin D1 protein expression and proliferation was further validated on an independent set of 23 MCLs.

Results:

MCLs expressed variable levels of cyclin D1 at both transcript and protein levels. Furthermore, D1L and D1S were the predominant transcripts in 69% and 31% of cases, respectively. While only 9% of cases with dominance of D1L had blastoid histology, 60% of the cases with dominance of the D1S had blastoid features. Furthermore, the levels of D1L showed direct correlation with cyclin D1 protein expression and Ki-67%. Among these cases, and in the independent set of MCLs (n = 40), the level of cyclin D1 protein expression directly correlated with Ki-67%.

Conclusions:

MCLs express variable levels of cyclin D1 transcripts and protein, and have variable proliferation (Ki-67%). Cases with dominance of D1S transcripts are more likely to be of blastoid morphology. There is correlation between D1L transcripts levels, cyclin D1 protein expression and Ki-67%.
Fine needle aspiration cytology of papillary lesions of the breast: how accurate is the diagnosis?
Authors: Tse G M K , Ma T K F , Lui P C W , Ng D C H , Yu A M C , Vong J S L , Niu Y , Chaiwun B , Lam W W M , Tan P H
Published: June 2008

Background:

Cytological diagnosis of mammary papillary lesions is difficult.

Aim:

To review the previous cytology diagnosis of 23 papillomas and 11 papillary carcinomas and specific cytological features that may assist in differentiating these entities.

Methods:

The cytology preparations were reviewed for: (i) overall cellularity; (ii) epithelial cell ball devoid of fibrovascular cores; (iii) background single cells; and (iv) papillary fragments and their morphology.

Results:

The overall diagnostic accuracy was 59%, atypical rate was 24%, and the error (combined false positive and negative) rate was 17%. For overall cellularity, 6, 14 and 3 cases of papillomas, and 6, 3 and 2 cases of papillary carcinomas showed low, moderate and high cellularity, respectively. Cell balls were present in mild to moderate number in 20 papillomas and 10 papillary carcinomas. The background single cells were absent, or present in low or moderate to high numbers in 7, 10 and 6 papillomas, and 3, 3 and 5 papillary carcinomas, respectively. Papillary fragments were absent, or present in small, moderate or large quantities in 9, 4, 8 and 2 papillomas, and 6, 3, 1 and 1 papillary carcinomas, respectively. There was no demonstrable quantitative difference between papilloma and papillary carcinoma for all these parameters. Qualitatively, the cell balls and single cells showed a higher degree of atypia in papillary carcinoma, and the papillary fragments were more elaborate and slender.

Conclusion:

Cytological diagnosis of papillary lesions shows a significant error rate with overlapping features. Cellular atypia and fragments with long and slender papillae with ramifying edges favour papillary carcinoma.
Do patients with low volume prostate cancer have prostate specific antigen recurrence following radical prostatectomy?
Authors: Furusato B , Rosner I L , Osborn D , Ali A , Srivastava S , Davis C J , Sesterhenn I A , McLeod D G
Published: June 2008

Aims:

The objective of this study was to determine the incidence of prostate specific antigen (PSA) relapse in patients with low volume prostate cancer following radical prostatectomy.

Methods:

Between 1993 and 2001, 50 of 717 patients had total tumour volumes of less than 0.5 cm3 following radical prostatectomy. Biochemical recurrence was defined as two consecutive values of serum PSA levels of 0.2 ng/ml or greater.

Results:

Median follow-up of the 50 patients was 58 months. In five of the 50 patients (10%), PSA recurrence was observed. All of these five cases had Gleason score of 3+3 (well and/or moderately differentiated), organ confined and surgical margin negative tumours. In three of the five cases, capsular incision resulted in benign glands extending into the surgical margin.

Conclusions:

Five of 50 cases had PSA failure. In three of the five patients, benign glands located in the margin could explain the “PSA recurrence”. However, in the other two patients, none of the pathological parameters correlated with measurable PSA levels. The explanation for their PSA failure is unclear.
Sediment roughness characteristics measured by broadband spectral analysis of acoustic echoes
Authors: Dunsiger A. D. , Cochrane N. A.
Published: June 2008

Acoustic signals scattered from the sea floor at nearly vertical incidence contain information about the physical properties of the sea floor. Signals may be decomposed into spatially coherent and spatially incoherent components the relative magnitudes of which are primarily a function of small scale bottom roughness. Experimental interping coherence functions derived on the basis of a linear system scattering model are shown to characterize contrasting sediment lithologies. The use of interping coherence as a sediment remote classification criterion and hence as a tracer of gross geotechnical properties is explored.
Effectiveness factor of Pt utilization in cathode catalyst layer of polymer electrolyte fuel cells
Authors: Xia Zetao , Wang Qianpu , Eikerling Michael , Liu Zhongsheng
Published: June 2008

In this work, we analyze effectiveness factors of Pt utilization in perfluorosulfonate ionomer (PFSI) bonded thin film cathode catalyst layers of polymer electrolyte fuel cells. We define the effectiveness factor of Pt utilization as the apparent rate of current conversion exhibited by a specific catalyst layer design divided by the ideal rate obtained if all Pt atoms were used equally in electrochemical reactions at the specified electrode overpotential and externally provided reactant concentrations. This definition includes statistical factors at all relevant scales as well as non-uniformities of reaction rate distributions under operation. Our model is based on the random composite agglomerated morphology of the catalyst layer. It accounts for the interplay of transport phenomena and electrochemical kinetics. At the mesoscopic scale, limited effectiveness of Pt utilization in agglomerates is mainly an electrostatic effect. We determined spatial distributions of effectiveness factors of agglomerates in the through-plane direction, and thereafter calculated overall effectiveness factors of the cathode catalyst layer. Our results show that small agglomerate radius, low operating current density, high operating temperature, and high oxygen partial pressure result in high effectiveness factors of Pt utilization. Finally, we compared PFSI-bonded thin film cathode catalyst layers with ultrathin two-phase cathode catalyst layers in terms of effectiveness factors. Including the surface to volume atom ratio of Pt nanoparticles, the two different types of structures exhibit similar effectiveness factors of Pt utilization, which are found to be distinctly below 10%.Key words: polymer electrolyte fuel cells, fuel cell modeling, cathode catalyst layer, Pt utilization, effectiveness factor.
Molecular identity of the late sodium current in adult dog cardiomyocytes identified by Nav1.5 antisense inhibition
Authors: Maltsev Victor A. , Kyle John W. , Mishra Sudhish , Undrovinas Abertas
Published: June 2008

Late Na+current ( INaL) is a major component of the action potential plateau in human and canine myocardium. Since INaLis increased in heart failure and ischemia, it represents a novel potential target for cardioprotection. However, the molecular identity of INaLremains unclear. We tested the hypothesis that the cardiac Na+channel isoform (Nav1.5) is a major contributor to INaLin adult dog ventricular cardiomyocytes (VCs). Cultured VCs were exposed to an antisense morpholino-based oligonucleotide (Nav1.5 asOligo) targeting the region around the start codon of Nav1.5 mRNA or a control nonsense oligonucleotide (nsOligo). Densities of both transient Na+current ( INaT) and INaL(both in pA/pF) were monitored by whole cell patch clamp. In HEK293 cells expressing Nav1.5 or Nav1.2, Nav1.5 asOligo specifically silenced functional expression of Nav1.5 (up to 60% of the initial INaT) but not Nav1.2. In both nsOligo-treated controls and untreated VCs, INaTand INaLremained unchanged for up to 5 days. However, both INaTand INaLdecreased exponentially with similar time courses (τ = 46 and 56 h, respectively) after VCs were treated with Nav1.5 asOligo without changes in 1) decay kinetics, 2) steady-state activation and inactivation, and 3) the ratio of INaLto INaT. Four days after exposure to Nav1.5 asOligo, INaTand INaLamounted to 68 ± 6% (mean ± SE; n = 20, P < 0.01) and 60 ± 7% ( n = 11, P < 0.018) of those in VCs treated by nsOligo, respectively. We conclude that in adult dog heart Nav1.5 sodium channels have a “functional half-life” of ∼35 h (0.69τ) and make a major contribution to INaL.
Nitrite reductase activity of hemoglobin as a systemic nitric oxide generator mechanism to detoxify plasma hemoglobin produced during hemolysis
Authors: Minneci Peter C. , Deans Katherine J. , Shiva Sruti , Zhi Huang , Banks Steven M. , Kern Steven , Natanson Charles , Solomon Steven B. , Gladwin Mark T.
Published: June 2008

Hemoglobin (Hb) potently inactivates the nitric oxide (NO) radical via a dioxygenation reaction forming nitrate (NO3−). This inactivation produces endothelial dysfunction during hemolytic conditions and may contribute to the vascular complications of Hb-based blood substitutes. Hb also functions as a nitrite (NO2−) reductase, converting nitrite into NO as it deoxygenates. We hypothesized that during intravascular hemolysis, nitrite infusions would limit the vasoconstrictive properties of plasma Hb. In a canine model of low- and high-intensity hypotonic intravascular hemolysis, we characterized hemodynamic responses to nitrite infusions. Hemolysis increased systemic and pulmonary arterial pressures and systemic vascular resistance. Hemolysis also inhibited NO-dependent pulmonary and systemic vasodilation by the NO donor sodium nitroprusside. Compared with nitroprusside, nitrite demonstrated unique effects by not only inhibiting hemolysis-associated vasoconstriction but also by potentiating vasodilation at plasma Hb concentrations of <25 μM. We also observed an interaction between plasma Hb levels and nitrite to augment nitroprusside-induced vasodilation of the pulmonary and systemic circulation. This nitrite reductase activity of Hb in vivo was recapitulated in vitro using a mitochondrial NO sensor system. Nitrite infusions may promote NO generation from Hb while maintaining oxygen delivery; this effect could be harnessed to treat hemolytic conditions and to detoxify Hb-based blood substitutes.
Integrin αvβ3acts downstream of insulin in normalization of interstitial fluid pressure in sepsis and in cell-mediated collagen gel contraction
Authors: Svendsen Øyvind Sverre , Lidén Åsa , Nedrebø Torbjørn , Rubin Kristofer , Reed Rolf K.
Published: June 2008

The administration of insulin is recommended to patients with severe sepsis and hyperglycemia. Previously, we demonstrated that insulin may have direct anti-inflammatory properties and counteracted fluid losses from the circulation by normalizing the interstitial fluid pressure (PIF). PIFis one of the Starling forces determining fluid flux over the capillary wall, and a lowered PIFis one of the driving forces in early edema formation in inflammatory reactions. Here we demonstrate that insulin restores a lipopolysaccharide (LPS)-lowered PIFvia a mechanism involving integrin αvβ3. In C57 black mice ( n = 6), LPS lowered PIFfrom −0.2 ± 0.2 to −1.6 ± 0.3 ( P < 0.05) and after insulin averaged −0.8 ± 0.2 mmHg ( P = 0.098 compared with after LPS). Corresponding values in wild-type BALB/c mice ( n = 5) were −0.8 ± 0.1, −2.1 ± 0.3 ( P < 0.05), and −0.8 ± 0.3 mmHg ( P < 0.05 compared with LPS) after insulin administration. In BALB/c integrin β3-deficient (β3−/−) mice ( n = 6), LPS lowered PIFfrom −0.1 ± 0.2 to −1.5 ± 0.3 mmHg ( P < 0.05). Insulin did not, however, restore PIFin these mice (averaged −1.7 ± 0.3 mmHg after insulin administration). Cell-mediated collagen gel contraction can serve as an in vitro model for in vivo measurements of PIF. Insulin induced αvβ3-integrin-dependent collagen gel contraction mediated by C2C12 cells. Our findings suggest a beneficiary effect of insulin for patients with sepsis with regard to the fluid balance, and this effect may in part be due to a normalization of PIFby a mechanism involving the integrin αvβ3.
Quantitative comparison of sarcomeric phosphoproteomes of neonatal and adult rat hearts
Authors: Yuan Chao , Sheng Quanhu , Tang Haixu , Li Yixue , Zeng Rong , Solaro R. John
Published: June 2008

Neonatal hearts respond to stress and function in an environment quite different from adult hearts. There is evidence that these functional differences not only reflect modifications in the abundance and isoforms of sarcomeric proteins but also in the modulation of sarcomeric protein phosphorylation. Yet our understanding of changes in sarcomeric protein phosphorylation in development is incomplete. In the experiments reported here, we first quantified the intact sarcomeric protein phosphorylation status between neonatal and adult rat hearts by employing comparative two-dimensional (2-D) gel electrophoresis in conjunction with phosphoprotein-specific staining. Subsequently, we measured phosphorylation changes at the peptide level by employing high-resolution linear ion trap-Fourier transform (LTQ-FT) mass spectrometry analysis of titanium dioxide-enriched phosphopeptides differentially labeled with16O/18O during in-gel digestion. We also employed Western blot analysis using phosphorylation site-specific antibodies to measure phosphorylation changes. Our data demonstrated the novel finding that phosphorylation levels of myosin-binding protein C (MyBP-C) at Ser295and Ser315as well as tropomyosin at Ser283increased, whereas phosphorylation levels of MyBP-C at Ser320and myosin light chain 2 at Ser15decreased in neonatal hearts compared with the same sites in adult hearts. Although our data highlight the significant challenges in understanding relations between protein phosphorylation and cardiac function, they do support the hypothesis that developmental changes in the modulation of protein are functionally significant and correlate with the prevailing physiological state.
Substrate uptake and metabolism are preserved in hypertrophic caveolin-3 knockout hearts
Authors: Augustus Ayanna S. , Buchanan Jonathan , Addya Sankar , Rengo Giuseppe , Pestell Richard G. , Fortina Paolo , Koch Walter J. , Bensadoun Andre , Abel E. Dale , Lisanti Michael P.
Published: June 2008

Caveolin-3 (Cav3), the primary protein component of caveolae in muscle cells, regulates numerous signaling pathways including insulin receptor signaling and facilitates free fatty acid (FA) uptake by interacting with several FA transport proteins. We previously reported that Cav3 knockout mice (Cav3KO) develop cardiac hypertrophy with diminished contractile function; however, the effects of Cav3 gene ablation on cardiac substrate utilization are unknown. The present study revealed that the uptake and oxidation of FAs and glucose were normal in hypertrophic Cav3KO hearts. Real-time PCR analysis revealed normal expression of lipid metabolism genes including FA translocase (CD36) and carnitine palmitoyl transferase-1 in Cav3KO hearts. Interestingly, myocardial cAMP content was significantly increased by 42%; however, this had no effect on PKA activity in Cav3KO hearts. Microarray expression analysis revealed a marked increase in the expression of genes involved in receptor trafficking to the plasma membrane, including Rab4a and the expression of WD repeat/FYVE domain containing proteins. We observed a fourfold increase in the expression of cellular retinol binding protein-III and a 3.5-fold increase in 17β-hydroxysteroid dehydrogenase type 11, a member of the short-chain dehydrogenase/reductase family involved in the biosynthesis and inactivation of steroid hormones. In summary, a loss of Cav3 in the heart leads to cardiac hypertrophy with normal substrate utilization. Moreover, a loss of Cav3 mRNA altered the expression of several genes not previously linked to cardiac growth and function. Thus we have identified a number of new target genes associated with the pathogenesis of cardiac hypertrophy.
The physiological basis of intracrine stem cell regulation
Authors: Re Richard N. , Cook Julia L.
Published: June 2008

Intracrine peptides and proteins participate in the regulation of adult and pleuripotential embryonic-like stem cells. Included among these factors are VEGF, dynorphin, the readthrough form of acetylcholinesterase, Oct3/4, Pdx-1, Pax-6, and high-mobility group protein B1, among others. In some cases, the establishment of intracrine feedback loops can be shown to be relevant to this regulation, consistent with previously proposed principles of intracrine action. Here the role of intracrines in stem cell regulation is reviewed, with particular attention to the intracrine regulation of cardiac stem cells. The reprogramming of cells to restore the pleuripotent phenotype and the possible role of stem/progenitor cells in neoplasia are also discussed.
GATA4 is a survival factor in adult cardiac myocytes but is not required for α1A-adrenergic receptor survival signaling
Authors: Huang Yuan , Wright Casey D. , Kobayashi Satoru , Healy Chastity L. , Elgethun Megan , Cypher Andrew , Liang Qiangrong , O'Connell Timothy D.
Published: June 2008

Recently, we defined an α1A-adrenergic receptor-ERK (α1A-AR-ERK) survival signaling pathway in adult cardiac myocytes. Previous studies in neonatal cardiac myocytes indicated that the cardiac-specific transcription factor GATA4 is a downstream mediator of α1-ERK signaling and that phosphorylation of GATA4 by ERK increases DNA binding and transcriptional activity. Therefore, we examined GATA4 as a potential downstream effector of α1A-ERK survival signaling in adult cardiac myocytes. We measured norepinephrine (NE)-induced cell death in cultured cardiac myocytes lacking α1-ARs (cultured from α1A/B-AR double-knockout mice, α1ABKO mice) that are susceptible to cell death induced by several proapoptotic stimuli, including NE. Our results show that overexpression of GATA4 is sufficient to protect α1ABKO cardiac myocytes from NE-induced cell death. However, we found that the α1A-subtype did not induce phosphorylation or increase the activity of GATA4 in adult mouse cardiac myocytes in culture or in vivo. Furthermore, we examined the effect of siRNA-mediated knockdown of GATA4 on α1A-survival signaling. In α1B-knockout cardiac myocytes, which express only the α1A-subtype and are protected from NE-induced cell death, GATA4 knockdown did not reverse α1A-survival signaling in response to NE. In summary, we found that GATA4 acted as a survival factor by preventing cell death in α1ABKO cardiac myocytes, but GATA4 was not activated by α1-AR stimulation and was not required for α1A-survival signaling in adult cardiac myocytes. This also identifies an important mechanistic difference in α1-signaling between adult and neonatal cardiac myocytes.
Inflammation, endothelial injury, and persistent pulmonary hypertension in heterozygous BMPR2-mutant mice
Authors: Song Yanli , Coleman Laura , Shi Jianru , Beppu Hideyuki , Sato Kaori , Walsh Kenneth , Loscalzo Joseph , Zhang Ying-Yi
Published: June 2008

Heterozygous bone morphogenetic protein receptor-II-knockout (BMPR2+/−) mice have a similar genetic trait like that in some idiopathic pulmonary arterial hypertension patients. To examine the effect of pulmonary endothelial injury in BMPR2+/−mice, we challenged the mice with two injections of monocrotaline combined with intratracheal instillation of replication-deficient adenovirus expressing 5-lipoxygenase (MCT+Ad5LO). After the challenge (1 wk), BMPR2+/−mice exhibited a doubling of right ventricular systolic pressure that was greater than that of wild-type mice and remained elevated for 3 wk before heart failure developed. Muscularization and thickening of small pulmonary arterioles was evident in the BMPR2+/−lungs at 2 wk after the challenge and became severe at 3 wk. Marked perivascular infiltration of T cells, B cells, and macrophages was associated with the remodeled vessels. Real-time PCR analysis showed that the expression of six endothelial cell markers in lung tissue was decreased to 20–40% of original levels at 1 wk after the challenge in both BMPR2+/−and wild-type mice and largely recovered in wild-type (50–80%) but not BMPR2+/−lungs (30–50%) at 3 wk after the challenge. Macrophage inflammatory protein-1α and fractalkine receptor expression doubled in BMPR2+/−compared with wild-type lungs. Expression of type I and type II BMP receptors, but not transforming growth factor-β receptors, in the challenged BMPR2+/−and wild-type lungs showed a similar pattern of expression as that of endothelial markers. Apoptotic responses at 1 wk after MCT and Ad5LO challenge were also significantly greater in the BMPR2+/−lungs than the wild-type lungs. These data show that BMPR2+/−mice are more sensitive to MCT+Ad5LO-induced pulmonary hypertension than wild-type mice. Greater endothelial injury and an enhanced inflammatory response could be the underlying causes of the sensitivity and may work in concert with BMPR2 heterozygosity to promote the development of persistent pulmonary hypertension.
Hydrogen peroxide decreases endothelial nitric oxide synthase promoter activity through the inhibition of AP-1 activity
Authors: Kumar Sanjiv , Sun Xutong , Wedgwood Stephen , Black Stephen M.
Published: June 2008

Previously, we have reported that endothelial nitric oxide synthase (eNOS) promoter activity is decreased in pulmonary arterial endothelial cells (PAECs) in response to hydrogen peroxide (H2O2). Thus the objective of this study was to identify the cis-element(s) and transcription factor(s) responsible for oxidant-mediated downregulation of the eNOS gene. Initial promoter experiments in PAECs treated with H2O2revealed a significant decrease in activity of a promoter fragment containing 840 bp of upstream sequence of the human eNOS gene fused to a luciferase reporter. However, a promoter construct containing only 640 bp of upstream sequence had a significantly attenuated response to H2O2challenge. As the 840-bp promoter construct had a putative binding site for the transcription factor activator protein-1 (AP-1) that was lacking in the 640-bp construct, we evaluated the effect of H2O2on promoter activity after mutation of the AP-1 binding sequence (TGAGTCA at −661 to TGAGTtg in the 840-bp construct). Similar to the results seen with the 640 bp, the AP-1 mutant promoter had a significantly attenuated response to H2O2. EMSA revealed decreased binding of AP-1 during H2O2treatment. Supershift analysis indicated that the AP-1 complex consisted of a c-Jun and FosB heterodimer. Furthermore, in vitro EMSA analysis indicated the c-Jun binding was significantly decreased after H2O2exposure. Using chromatin immunoprecipitation analysis, we demonstrated decreased binding of AP-1 to the eNOS promoter in vivo in response to H2O2. These data suggest a role of decreased AP-1 binding likely through c-Jun in the H2O2-mediated decrease in eNOS promoter activity.
Peroxisome proliferator-activated receptor-γ in cystic fibrosis lung epithelium
Authors: Perez Aura , van Heeckeren Anna M. , Nichols David , Gupta Sanhita , Eastman Jean F. , Davis Pamela B.
Published: June 2008

The pathophysiology of cystic fibrosis (CF) inflammatory lung disease is not well understood. CF airway epithelial cells respond to inflammatory stimuli with increased production of proinflammatory cytokines as a result of increased NF-κB activation. Peroxisome proliferator-activated receptor-γ (PPARγ) inhibits NF-κB activity and is reported to be reduced in CF. If PPARγ participates in regulatory dysfunction in the CF lung, perhaps PPARγ ligands might be useful therapeutically. Cell models of CF airway epithelium were used to evaluate PPARγ expression and binding to NF-κB at basal and under conditions of inflammatory stimulation by Pseudomonas aeruginosa or TNFα/IL-1β. An animal model of CF was used to evaluate the potential of PPARγ agonists as therapeutic agents in vivo. In vitro, PPARγ agonists reduced IL-8 and MMP-9 release from airway epithelial cells in response to PAO1 or TNFα/IL-1β stimulation. Less NF-κB bound to PPARγ in CF than normal cells, in two different assays; PPARγ agonists abrogated this reduction. PPARγ bound less to its target DNA sequence in CF cells. To test the importance of the reported PPARγ inactivation by phosphorylation, we observed that inhibitors of ERK, but not JNK, were synergistic with PPARγ agonists in reducing IL-8 secretion. In vivo, administration of PPARγ agonists reduced airway inflammation in response to acute infection with P. aeruginosa in CF, but not wild-type, mice. In summary, PPARγ inhibits the inflammatory response in CF, at least in part by interaction with NF-κB in airway epithelial cells. PPARγ agonists may be therapeutic in CF.
Mdm2 and Mdm4 Loss Regulates Distinct p53 Activities
Authors: Barboza Juan A. , Iwakuma Tomoo , Terzian Tamara , El-Naggar Adel K. , Lozano Guillermina
Published: June 2008

Mutational inactivation of p53 is a hallmark of most human tumors. Loss of p53 function also occurs by overexpression of negative regulators such as MDM2 and MDM4. Deletion of Mdm2 or Mdm4 in mice results in p53-dependent embryo lethality due to constitutive p53 activity. However, Mdm2−/− and Mdm4−/− embryos display divergent phenotypes, suggesting that Mdm2 and Mdm4 exert distinct control over p53. To explore the interaction between Mdm2 and Mdm4 in p53 regulation, we first generated mice and cells that are triple null for p53, Mdm2, and Mdm4. These mice had identical survival curves and tumor spectrum as p53−/− mice, substantiating the principal role of Mdm2 and Mdm4 as negative p53 regulators. We next generated mouse embryo fibroblasts null for p53 with deletions of Mdm2, Mdm4, or both; introduced a retrovirus expressing a temperature-sensitive p53 mutant, p53A135V; and examined p53 stability and activity. In this system, p53 activated distinct target genes, leading to apoptosis in cells lacking Mdm2 and a cell cycle arrest in cells lacking Mdm4. Cells lacking both Mdm2 and Mdm4 had a stable p53 that initiated apoptosis similar to Mdm2-null cells. Additionally, stabilization of p53 in cells lacking Mdm4 with the Mdm2 antagonist nutlin-3 was sufficient to induce a cell death response. These data further differentiate the roles of Mdm2 and Mdm4 in the regulation of p53 activities. (Mol Cancer Res 2008;6(6):947–54)
Tumor Environment Dictates Medulloblastoma Cancer Stem Cell Expression and Invasive Phenotype
Authors: Annabi Borhane , Rojas-Sutterlin Shanti , Laflamme Carl , Lachambre Marie-Paule , Rolland Yannève , Sartelet Hervé , Béliveau Richard
Published: June 2008

The neural precursor surface marker CD133 is thought to be enriched in brain cancer stem cells and in radioresistant DAOY medulloblastoma-derived tumor cells. Given that membrane type-1 matrix metalloproteinase (MT1-MMP) expression is a hallmark of highly invasive, radioresistant, and hypoxic brain tumor cells, we sought to determine whether MT1-MMP and other MMPs could regulate the invasive phenotype of CD133(+) DAOY cells. We found that when DAOY medulloblastoma or U87 glioblastoma cells were implanted in nude mice, only those cells specifically implanted in the brain environment generated CD133(+) brain tumors. Vascular endothelial growth factor and basic fibroblast growth factor gene expression increases in correlation with CD133 expression in those tumors. When DAOY cultures were induced to generate in vitro neurosphere-like cells, gene expression of CD133, MT1-MMP, MMP-9, and MDR-1 was induced and correlated with an increase in neurosphere invasiveness. Specific small interfering RNA gene silencing of either MT1-MMP or MMP-9 reduced the capacity of the DAOY monolayers to generate neurospheres and concomitantly abrogated their invasive capacity. On the other hand, overexpression of MT1-MMP in DAOY triggered neurosphere-like formation which was further amplified when cells were cultured in neurosphere medium. Collectively, we show that both MT1-MMP and MMP-9 contribute to the invasive phenotype during CD133(+) neurosphere-like formation in medulloblastoma cells. Increases in MMP-9 may contribute to the opening of the blood-brain barrier, whereas increased MT1-MMP would promote brain tumor infiltration. Our study suggests that MMP-9 or MT1-MMP targeting may reduce the formation of brain tumor stem cells. (Mol Cancer Res 2008;6(6):907–16)
The Leaf Fell (the Leaf): The Online Processing of Unaccusatives
Authors: Friedmann Naama , Taranto Gina , Shapiro Lewis P. , Swinney David
Published: June 2008

According to the Unaccusative Hypothesis, unaccusative subjects are base-generated in object position and move to subject position. We examined this hypothesis using the cross-modal lexical priming technique, which tests whether and when an antecedent is reactivated during the online processing of a sentence. We compared sentences containing unergative verbs with sentences containing unaccusatives, both alternating and nonalternating, and found that subjects of unaccusatives reactivate after the verb, while subjects of unergatives do not. Alternating unaccusatives showed a mixed pattern of reactivation. The research directly supports the Unaccusative Hypothesis.
Services - Led Growth in Thailand, 1956-1970
Authors: Nartsupha Chatthip
Published: June 2008

The purpose of this short paper is to describe briefly the peculiar pattern of Thailand's economic growth during the last 15 years. The paper will be divided into three parts. The first part will briefly describe the pattern of Thailand's economic development from 1956 to 1970. The second part will show the impact of this particular pattern of development on Thailand's balance of payments. The third part will suggest some other possible effects of this development pattern.
Ethical Systems and Economic Development
Authors: Jonassen Christen T.
Published: June 2008

The possible role of ethical systems in accounting for the wide economic and social gap betweeri developed and underdeveloped countries is examined in the light of the Weberian thesis on the relationship between the Protestant Ethic, capitalism and economic development. Materials from various cultures seem to support Weber, but suggest that the crucial factor in economic development is not necessarily THE PROTESTANT ETHIC, but any ethical system which produces the following results: rational attitudes toward experiences, personal responsibility, a disciplined work force, and worldly asceticism. An ethic which encourages underconsumption and deferred gratification and which creates social motivations and legitimization for worldly work and economic enterprise seems to facilitate economic development in the takeoff period.
Hindu Mediumship in Singapore 1
Authors: Babb Lawrence A.
Published: June 2008

It is'a commonplace to ethnographers of Singapore that the island has provided a fertile environment for the flourishing of Chinese spirit mediumship. This pheonomenon has been well-described elsewhere (see Elliot). It is perhaps less well known that medium- . ship constitutes an extremely important element in the religious life of Singapore's Indian minority. It is with Hindu mediumship in Singapore that the present essay is concerned. My aims are two: to describe some of the most salient features of the institution, and to show how it fits into broader patterns of Hindu religious belief and practice as seen in Singapore.. ! I hope to show that Hindu mediumship is an important component of a more inclusive religious system, a component which functions to mediate between the highly particularized religious interests of individuals and the abstract conceptions of divine power which seem to underlie the religious system as a whole. The data upon which the study is based were drawn from formal and casual interviews with many informants, and from focussed investigations of the activities of five different Hindu mediums. This sample is not random. Hindu mediums operate in relative obscurity and there seems to be no way to search out a sample in a systematic way. I heard of each medium from a friend or acquaintance who thought I might be interested. and in this respect my initial encounters with them were not unlike their initial encounters with their normal clients. In most cases my relationship with the medium became a dual one. I identified myself as a researcher interested in Hinduism, and I interviewed mediums, clients and hangers-on in this spirit. But at the same time there was often what seemed to be an inevitable drift from the role of investigator to that of client, the latter being the only legitimate niche which the situation seemed to be able to provide for my presence. Accordingly, I participated in ceremonies and, with other clients, sought the counsel of the gods through their human representatives.
THE POWER ELITE IN THAILAND: A General Survey with a Focus on the Civil Bureaucrats
Authors: Dhiravegin Likhit
Published: June 2008

At the macro level, we have given a systemic picture of the elite structure of Thai society. We have argued that the most three powerful groups of elite are the military, the civil bureaucratic elite and the business elite. Among these three groups of elite, an alliance has been developed. The business elite have sought political protection from the military and the bureaucratic elite while the latter. two groups obtain economic benefit. Below the three groups of elite lie the people's representatives; the student leaders, the labour leaders and the peasant organizers. We have argued that the people's representatives are on the border line because the democratice structure has not yet become institutionalized. And this is true also for the student leaders, the labour leaders and the peasant organizers. At best, they are at the early stage of getting organized and thus they are classified as interest groups who are becoming more and more active in politics. At the micro level, we have presented an analysis of the general characteristics of the civil bureaucratic elite of Thailand. We have found from our discussion that more than two-fifths (41%) of the elite in our study have fathers in government service; 31.44% are from families of business; only 5.6% are from a peasant background. There ia an inbreeding in the government service career. We have hypothesized that family background plays a crucial role in an individual's educational achievement which, in turn, leads to his social mobility. From our study, we have found that social mobility