bluuebunny/crossref_metadata_2025 huggingface dataset:
The mechanism of improved sodium homeostasis of low-dose losartan in preascitic cirrhosis
Authors: Wong Florence , Liu Peter , Blendis Laurence
Published: March 2004

Renal sodium retention on standing is one aspect of the abnormal renal sodium handling in preascitic, well-compensated patients with cirrhosis. Recently, it has been shown that low doses (7.5 mg) of the angiotensin II (Ang II) receptor antagonist, losartan, can reverse renal sodium retention on high, 200-mmol sodium/d diet in these patients and restore them to sodium balance. Therefore, the effect of 7.5 mg of losartan on sodium excretion, when changing from supine to erect posture for 2 hours, was examined in 10 well-compensated patients with cirrhosis and 9 age- and sex-matched controls on the same sodium diet, under strictly controlled metabolic conditions. In contrast to control subjects, in whom sodium excretion was unaffected, single 7.5-mg doses of losartan again restored the preascitic patients with cirrhosis to sodium balance. In addition, it blunted the fall in erect posture- induced renal sodium excretion by a reduction in proximal and distal tubular reabsorption of sodium. These changes occurred without any significant changes in blood volumes, systemic and renal hemodynamics, or glomerular filtration rate (GFR) and filtered sodium load compared with controls, and despite activation of the systemic renin-angiotensin-aldosterone system, which was still within normal levels. In conclusion, the beneficial natriuretic effects of low-dose losartan on erect posture - induced sodium retention in preascitic cirrhosis supports the suggestion that the pathophysiology of sodium retention in preascites is in part caused by an intrarenal tubular effect of Ang II in that posture.
Diminished hepatic expression of the HNF-6 transcription factor during bile duct obstruction
Authors: Holterman Ai-Xuan L. , Tan Yongjun , Kim Wooram , Yoo Kyung W. , Costa Robert H.
Published: March 2004

Hepatocyte nuclear factor 6 (HNF-6) is a member of the one cut family of transcription factors and potentially regulates expression of numerous target genes important for hepatocyte function. In the liver, HNF-6 is expressed not only in hepatocytes, but also in biliary epithelial cells (BEC). To evaluate the in vivo function of HNF-6, we examined the hepatic expression pattern of HNF-6 messenger RNA (mRNA) and protein after bile duct ligation (BDL)-mediated liver injury. We found that HNF-6 protein levels in BEC and hepatocytes were diminished within 15 hours of BDL injury and remained suppressed through the fifth day of injury. The onset of BEC proliferation in response to bile duct obstruction was associated with diminished HNF-6 protein levels. To maintain hepatic HNF-6 protein levels during BDL liver injury, we used mouse tail vein injections with recombinant adenovirus expressing HNF-6 complementary DNA (cDNA) (AdH6). We found that maintaining hepatic HNF-6 levels with AdH6 infection resulted in significant decreases in BEC proliferation at 15 and 24 hours after biliary obstruction compared with adenovirus control. Our results showed that HNF-6 expression is diminished in BEC and hepatocytes and that maintaining hepatic HNF-6 expression hinders the normal biliary proliferative response to bile duct injury. This suggests that diminished hepatic HNF-6 levels are required for repair in response to biliary injury and that it regulates expression of genes that possess differentiation-specific function that are inhibitory to proliferation. In conclusion, we propose a biologic role for diminished HNF-6 protein levels in bile duct disease.
Nonsurgical treatment of hepatocellular carcinoma
Authors: Llovet Josep M. , Sala Margarita , Bruix Jordi
Published: March 2004

Key Points

1. Outcome from nonsurgical treatment is directly related to stage of hepatocellular cancer (HCC) and degree of liver function impairment.

2. Ablative percutaneous procedures, such as alcohol injection or radiofrequency thermal therapy, are most effective in the destruction of solitary tumors of 3 cm or less.

3. In most cases, nonsurgical treatments are not curative, but may slow tumor progression and can provide palliation.

4. Arterial embolization or chemoembolization has an antitumor effect, but it has not been shown to affect patient outcome.

5. Radiation therapy, chemotherapy, hormonal manipulation, and interferon have not been consistently effective in HCC.

6. Ablative procedures, embolization, and systemic chemotherapy should be avoided in patients with advanced cirrhosis.
An efficacy and cost-effectiveness analysis of combination hepatitis B immune globulin and lamivudine to prevent recurrent hepatitis B after orthotopic liver transplantation compared with hepatitis B immune globulin monotherapy
Authors: Han Steven-Huy B. , Ofman Joshua , Holt Curtis , King Kevin , Kunder Gregg , Chen Pauline , Dawson Sherfield , Goldstein Leonard , Yersiz Hasan , Farmer Douglas G. , Ghobrial Rafik M. , Busuttil Ronald W. , Martin Paul
Published: March 2004

Orthotopic liver transplantation (OLT) for hepatitis B virus (HBV) infection was limited until recently by poor graft and patient outcomes caused by recurrent HBV. Long-term immunoprophylaxis with hepatitis B immune globulin (HBIG) dramatically improved post-OLT survival, but recurrent HBV still occurred in up to 36% of the recipients. More recently, combination HBIG and lamivudine has been shown to effectively prevent HBV recurrence in patients post-OLT. The aim of the current study is to determine long-term outcome and cost-effectiveness of using combination HBIG and lamivudine compared with HBIG monotherapy in patients who undergo OLT for HBV. A retrospective chart review identified 59 patients administered combination HBIG and lamivudine and 12 patients administered HBIG monotherapy as primary prophylaxis against recurrent HBV. Lamivudine, 150 mg/d, was administered orally indefinitely. HBIG was administered under a standard protocol (10,000 IU intravenously during the anhepatic phase, then 10,000 IU/d intravenously for 7 days, then 10,000 IU intravenously monthly) indefinitely. A decision-analysis model was developed to evaluate the potential economic impact of prophylaxis against HBV with combination therapy compared with monotherapy. Recurrent HBV was defined as the reappearance of hepatitis B surface antigen (HBsAg) after its initial disappearance post-OLT. In the combination-therapy group, no patient redeveloped serum HBsAg or HBV DNA during mean follow-ups of 459 and 416 days, respectively. In the monotherapy group, 3 patients (25%) had reappearance of HBsAg in serum during a mean follow-up of 663 days. Combination therapy resulted in a dominant, cost-effective strategy with an average cost-effectiveness ratio of $252,111/recurrence prevented compared with $362,570/recurrence prevented in the monotherapy strategy. Combination prophylaxis with HBIG and lamivudine is highly effective in preventing recurrent HBV, may protect against the emergence of resistant mutants, and is significantly more cost-effective than HBIG monotherapy with its associated rate of recurrent HBV.
Posttransplantation prevention and treatment of recurrent hepatitis B
Authors: Rizzetto Mario , Marzano Alfredo
Published: March 2004

Key Points

Prophylaxis of Recurrent Hepatitis B

1. Although standard prophylaxis with antibody to hepatitis B surface antigen immunoglobulins (HBIG) is effective, it is difficult to administer and must be administered indefinitely.

2. Preemptive therapy with lamivudine reduces the early risk for recurrence after transplantation, but maintenance with either famciclovir or lamivudine has been ineffective in sustaining remission.

3. The combination of preemptive lamivudine with HBIG prophylaxis may be the most effective treatment to prevent hepatitis B virus reinfection.

Treatment of Recurrent Hepatitis B

1. Interferon-α, ganciclovir, and famciclovir have not been helpful.

2. Lamivudine appears promising, but its long-term efficacy is unproven; in immunosuppressed transplant recipients, the rate of emergence of YMDD mutants is high and accelerated, and their emergence is aggravated by consistent liver morbidity.
Liver transplantation for polycystic liver disease
Authors: Pirenne Jacques , Aerts Raymond , Yoong Kim , Gunson Bridget , Koshiba Takaaki , Fourneau Inge , Mayer David , Buckels John , Mirza Darius , Roskams Tania , Elias Elwin , Nevens Frederik , Fevery Johan , McMaster Paul
Published: March 2004

Polycystic liver disease (PLD) may provoke massive hepatomegaly and severe physical and social handicaps. Data on orthotopic liver transplantation (OLT) for PLD are rare and conflicting. Conservative surgery (resection or fenestration) is indicated for large single cysts, but its value for small diffuse cysts is questionable. In addition, conservative surgery is not devoid of morbidity and mortality. OLT offers the prospect of a fully curative treatment, but controversy remains because those patients usually have preserved liver function. Thus, we reviewed our experience with OLT for PLD. Sixteen adult women underwent OLT for small diffuse PLD between 1990 and 1999. Mean age was 45 years (range, 34 to 56 years). Fourteen patients had combined liver and kidney cystic disease, but only 1 patient required combined liver and kidney transplantation, whereas 13 patients underwent OLT alone. Two patients had isolated PLD. Indications for transplantation were massive hepatomegaly causing physical handicaps (n = 16), social handicaps (n = 16), malnutrition (n = 4), and cholestasis and/or portal hypertension (n = 5). OLT caused no technical difficulty in 15 of 16 patients (surgery duration, 6.8 hours; range, 5 to 8 hours), with blood transfusions of 7.9 units (range, 0 to 22 units). One patient who underwent attempted liver-mass reduction pre-OLT died of bleeding and pulmonary emboli. Native liver weight was 10 to 20 kg. Posttransplantation immunosuppression consisted of cyclosporine or FK506, azathioprine, and steroids (discontinued at 3 months). Morbidity included biliary stricture (2 patients), revision for bleeding and hepatitis (1 patient), pneumothorax and subphrenic collection (1 patient), and tracheostomy (1 patient). One patient died of lung cancer 6 years posttransplantation. Both patient and graft survival rates are 87.5% (follow-up, 3 months to 9 years). Of 15 patients who underwent OLT alone, only 1 patient needed a kidney transplant 4 years after OLT. Kidney function has remained satisfactory in the other patients despite the use of cyclosporine or FK506 (last follow-up creatinine level, 1.55 mg/dL; range, 0.80 to 2.85 mg/dL). OLT had a dramatic impact on daily quality of life, enabling these patients to go back to a fully active life style. OLT offers the chance of a definitive treatment in patients with extensive, small, diffuse PLD that has evolved into severely handicapping hepatomegaly. In contrast to previous studies, combined liver and kidney transplantation is rarely needed. Patient symptoms and chances of definitive palliation offered by OLT must be balanced against the risks of transplantation and lifelong commitment to immunosuppression.
Development of nonalcoholic fatty liver disease after orthotopic liver transplantation for cryptogenic cirrhosis
Authors: Contos Melissa J. , Cales Wendy , Sterling Richard K. , Luketic Velimir A. , Shiffman Mitchell L. , Mills Scott A. , Fisher Robert A. , Ham John , Sanyal Arun J.
Published: March 2004

Many subjects with cryptogenic cirrhosis have underlying nonalcoholic steatohepatitis (NASH). The natural history of NASH-related cryptogenic cirrhosis after orthotopic liver transplantation (OLT) is not well defined. A primarily retrospective study of patients with the clinical histological phenotype of NASH-related cirrhosis undergoing OLT was performed. Data were compared with 2 sets of age- and weight-matched controls with (1) primary biliary cirrhosis or primary sclerosing cholangitis or (2) alcoholic liver disease. After OLT, all patients were managed by a standard immunosuppressive protocol. Liver biopsies were performed at 6 and 12 months after OLT and at 1- to 2-year intervals thereafter, as well as when liver enzyme levels were elevated enough to warrant diagnostic biopsy. Twenty-seven subjects with cryptogenic cirrhosis and a clinical histological phenotype of NASH and 3 patients with a long-standing diagnosis of NASH before OLT were included. The 30-day perioperative mortality was 1 in 30 patients. During a median follow-up of 3.5 ± 2.7 years, 2 additional patients died of sepsis. There was a time-dependent increase in the risk for allograft steatosis that approached 100% by 5 years compared with only an approximately 25% incidence of steatosis in the control groups (P < .009, log-rank test). On multivariate analysis, only the cumulative steroid dose correlated with time to development of allograft steatosis. Three patients developed histological progression from hepatic steatosis to steatohepatitis. Of these, 1 patient developed progressive fibrosis. Four patients experienced at least 1 episode of acute cellular rejection; however, no patient developed chronic rejection or graft failure. In conclusion, nonalcoholic fatty liver disease occurs frequently after OLT in patients with the phenotype of NASH-related cirrhosis. Despite the frequent histological recurrence of disease, clinical outcomes are similar to those of other groups of patients undergoing OLT. (Liver Transpl 2001;7:363-373.)
Liver transplantation using sirolimus and minimal corticosteroids (3-day taper)
Authors: Trotter James F. , Wachs Michael , Bak T. , Trouillot Thomas , Stolpman Nancy , Everson Gregory T. , Kam Igal
Published: March 2004

At our center, we have performed liver transplantation since 1995 with a rapid-taper steroid protocol (weaning steroids by day 14 posttransplantation). Beginning in 2000, we further reduced the use of corticosteroids to 3 days and added sirolimus to our immunosuppressive regimen. We report our experience with 39 patients who underwent liver transplantation with either tacrolimus or cyclosporin A (Neoral; Novartis Pharmaceuticals Corp., Summit, NJ) and sirolimus, with a 3-day tapered dose of corticosteroids. Thirty-two patients received a cadaveric graft and 7 patients received a right hepatic lobe from a living donor. All patients initially were administered either tacrolimus (0.1 mg/kg/d) or cyclosporin A (10 mg/kg/d) and sirolimus (6 mg/d for 1 day, followed by 2 mg/d), in addition to methylprednisolone on the first 3 days (1, 0.5, and 0.5 g/d) after transplantation. Patients were administered corticosteroids for presumptive or biopsy-proven evidence of acute cellular rejection (methylprednisolone, 1, 0.5, and 0.5 g on 3 successive days). Seventeen patients were administered tacrolimus and 22 patients were administered cyclosporin A. Six patients were excluded from analysis because they were administered sirolimus for less than 2 weeks. Mean duration of follow-up was 124 days. Patient survival was 36 of 39 patients (92%), and graft survival was 35 of 39 grafts (89%). Ten of 33 patients (30%) experienced 12 episodes of rejection (7 biopsy proven, 5 presumptive) compared with 70% in historical controls (P < .01). OKT3 was required in 1 of 33 patients (3%) compared with 37% in controls (P < .01). Twenty-six of 33 patients (79%) were not administered prednisone, and 7 of 33 patients (21%) were administered prednisone for reasons other than rejection. Posttransplantation, there was no significant change in values for creatinine, glucose, aspartate aminotransferase, bilirubin, cholesterol, and white blood cell counts. Platelet counts were significantly reduced, and hematocrits were significantly elevated (P < .05). Liver transplantation may be successfully performed with minimal use of corticosteroids by using sirolimus and either tacrolimus or cyclosporin A. Despite the absence of prednisone from our immunosuppressive protocol, the incidence of rejection and OKT3 use was lower than in historical controls. Patient and graft survival rates were identical to those of historical controls. The findings in this report will serve as the basis for a formal trial evaluating the efficacy of sirolimus in liver transplantation.
Delayed recurrence of hepatocellular carcinoma after liver transplantation: Detection of origin by chromosomal analysis
Authors: Al-Joundi Tammam , Gibson Sandra , Brunt Elizabeth M. , Shakil Obaid , Lee Randall S. , Di Bisceglie Adrian M.
Published: March 2004

We report on a 41-year-old man undergoing liver transplantation for chronic hepatitis C who presented 26 months later with hepatocellular carcinoma. No evidence of hepatocellular carcinoma could be found in the native liver, although features of small cell dysplasia were prominent. Although he had recurrent hepatitis C, the transplanted liver was not cirrhotic. Chromosomal analysis was used to resolve whether this was a de novo tumor or a recurrence of an unsuspected tumor present at the time of transplantation. This male patient had received a liver from a female donor, and in situ hybridization for the Y chromosome showed reactivity in the tumor but not in surrounding nontumorous liver. Thus, this is an example of the use of chromosomal analysis to resolve the origin of a tumor occurring in the transplant setting.
Intracerebral hemorrhage after liver transplantation
Authors: Wang Wei-Lin , Yang Zhen-Fan , Lo Chung-Mau , Liu Chi-Leung , Fan Sheung-Tat
Published: March 2004

We report 5 patients with intracerebral hemorrhage after orthotopic liver transplantation (OLT) and identify the possible risk factors. Between November 1991 and April 1999, 75 adult patients received 77 orthotopic liver transplants at Queen Mary Hospital, Hong Kong. Five patients (6.5%) developed intracerebral hemorrhage postoperatively. Clinical and laboratory data were reviewed, and potential risk factors were analyzed. The 5 patients developed intracerebral hemorrhage within 40 days (range, 1 to 37 days; median, 4 days) after OLT. The mortality rate was 80% (4 of 5 patients). The intraoperative blood transfusion volume (median, 17,200 mL; range, 15,750 to 30,360 mL) administered to patients who developed intracerebral hemorrhage postoperatively was significantly greater than that (median, 6,990 mL; range, 1,840 to 22,680 mL) for patients without the complication (P= .0008). Massive intraoperative transfusion (>15,000 mL) was required in all 5 patients (100%) with intracerebral hemorrhage but only 9 of 72 patients (12.5%) in the other group (P= .0001). Four of 5 patients (80%) with intracerebral hemorrhage had intraoperative hypotension compared with 7 of 72 patients (9.7%) in the other group (P= .001). No significant difference was found in age, prothrombin time (PT), activated partial thromboplastin time (APTT), incidence of hypertension, bleeding at extracerebral sites, cyclosporine A neurotoxicity, thrombocytopenia, hemodialysis, and sepsis between the patients with and without intracerebral bleeding. However, the median cumulative score of coagulation parameters (PT, APTT, platelet count) was significantly greater in the group with than without intracerebral bleeding (median score, 3 v1; P= .023). Intracerebral hemorrhage is 1 of the most disastrous complications after OLT. Intraoperative hypotension, massive intraoperative transfusion, and coagulopathy may be correlated with this complication.
Spatial chaos of trapped Bose–Einstein condensate in one-dimensional weak optical lattice potential
Authors: Chong Guishu , Hai Wenhua , Xie Qiongtao
Published: March 2004

The spatially chaotic attractor in an elongated cloud of Bose–Einstein condensed atoms perturbed by a weak optical lattice potential is studied. The analytical insolvability and numerical incomputability of the atomic number density are revealed by a perturbed solution that illustrates the unpredictability of the deterministic chaos. Although this could lead the nonphysical explosion and unboundedness to the numerical solution, the theoretical analysis offers a criterion to avoid them. Moreover, the velocity field is investigated that exhibits the superfluid property of the chaotic system.
Calcium signaling in chemorepellant Slit2-dependent regulation of neuronal migration
Authors: Xu Hua-tai , Yuan Xiao-bing , Guan Chen-bing , Duan Shumin , Wu Chien-ping , Feng Linyin
Published: March 2004

Migration of neuronal precursor cells in the developing brain is guided by extracellular cues, but intracellular signaling processes underlying the guidance of neuronal migration are largely unknown. By examining the migration of cerebellar granule neurons along the surface of cocultured astroglial cells, we found that an extracellular gradient of Slit2, a chemorepellant for neuronal migration
in vivo
, caused a reversal in the direction of migration without affecting the migration speed. A Slit2 gradient elevated the intracellular concentration of Ca
2+
, probably due to calcium release from the internal store, led to a reversal of the preexisting asymmetric intracellular Ca
2+
distribution in the soma of migrating neurons, and this reversal was closely related with its action of reversing the migrating direction. Asymmetric Ca
2+
distribution in the soma was both necessary and sufficient for directing neuronal migration. These results have demonstrated an important role for Ca
2+
in mediating neuronal responses to Slit2 and suggest a general mechanism for neuronal guidance.
The ecological and evolutionary consequences of sperm chemoattraction
Authors: Riffell Jeffrey A. , Krug Patrick J. , Zimmer Richard K.
Published: March 2004

Chemical communication between sperm and egg is a critical factor mediating sexual reproduction. Sperm attractants may be significant evolutionarily for maintaining species barriers, and important ecologically for increasing gamete encounters. Still unresolved, however, are the functional consequences of these dissolved signal molecules. Here, we provide experimental evidence that sperm chemoattraction directly affects the magnitude of fertilization success. The recent discovery of
l
-tryptophan as a potent attractant to red abalone (
Haliotis rufescens
) sperm affords the opportunity to quantify how navigation affects gamete interactions. Sperm behavioral responses to manipulations of the natural tryptophan gradient around individual eggs reveals that both chemotaxis and chemokinesis significantly promote contacts. Our results show further that attractant release by means of diffusion effectively doubles the target size of red abalone eggs, which in turn significantly increases fertilization success. Although long theorized as potential barriers to hybridization, species-specific sperm attractants in red and green (
Haliotis fulgens
) abalone are only minor contributors to maintaining reproductive isolation. Because abalone typically live in dense, multispecies aggregations, chemically mediated navigation would prevent sperm from pointlessly tracking heterospecific eggs. Thus, even though reproductive isolation fundamentally resides at the level of membrane recognition proteins, species-specific sperm attractants may evolve to locate the right target within mixed gamete suspensions of closely related species.
Pressure-dissociable reversible assembly of intrinsically denatured lysozyme is a precursor for amyloid fibrils
Authors: Niraula Tara N. , Konno Takashi , Li Hua , Yamada Hiroaki , Akasaka Kazuyuki , Tachibana Hideki
Published: March 2004

Although a diversity of proteins is known to form amyloid fibers, their common mechanisms are not clear. Here, we show that an intrinsically unfolded protein (U), represented by a disulfide-deficient variant of hen lysozyme with no tertiary structure, forms an amyloid-like fibril after prolonged incubation. Using variable pressure NMR along with sedimentation velocity, circular dichroism, and fluorescence measurements, we show that, before the fibril formation, the protein forms a pressure-dissociable, soluble assemblage (

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\begin{equation*}{\mathrm{U}}_{{\mathrm{n}}}^{^{\prime}}\end{equation*}\end{document}

) with a sedimentation coefficient of 17 S and a rich intermolecular β-sheet structure. The reversible assemblage is characterized with a Gibbs energy for association of –23.3 ± 0.8 kJ·mol
–1
and a volume increase of 52.7 ± 11.3 ml·mol
–1
per monomer unit, and involves preferential interaction of hydrophobic residues in the initial association step. These results indicate that amyloid fibril formation can proceed from an intrinsically denatured protein and suggest a scheme

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as a common mechanism of fibril formation in amyloidogenic proteins, where two-way arrows represent reversible processes, one-way arrow represents an irreversible process, and N, U, and

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represent, respectively, the native conformer, the unfolded monomeric conformer, and the soluble assemblage of unfolded conformers.
Transplantation of skeletal myoblasts secreting an IL-1 inhibitor modulates adverse remodeling in infarcted murine myocardium
Authors: Murtuza Bari , Suzuki Ken , Bou-Gharios George , Beauchamp Jonathan R. , Smolenski Ryszard T. , Partridge Terence A. , Yacoub Magdi H.
Published: March 2004

After myocardial infarction (MI), adverse remodeling with left ventricular (LV) dilatation is a major determinant of poor outcome. Skeletal myoblast (SkM) implantation improves cardiac function post-MI, although the mechanism is unclear. IL-1 influences post-MI hypertrophy and collagen turnover and is implicated in SkM death after grafting. We hypothesized that SkM expressing secretory IL-1 receptor antagonist (sIL-1ra) at MI border zones would specifically attenuate adverse remodeling and exhibit improved graft cell number. Stable murine male SkM lines (5 × 105cells), expressing or nonexpressing (cont) for sIL-1ra, were implanted into infarct border zones of female nude mice immediately after left coronary artery occlusion. LV ejection fraction (LVEF), end-diastolic diameter, and transmitral peak early/late (E/A) flow velocity ratio were determined by echocardiography. Cardiac myocyte hypertrophy and fibrosis were assessed by morphometry, picrosirius red staining, and hydroxyproline assay. At 3 weeks, cont-SkM-engrafted hearts showed reduced hypertrophy, improved LVEF (55.7 ± 1.2% vs. MI-only: 40.3 ± 2.9%), and preserved E/A ratios. sIL-1ra-SkM implantation enhanced these effects (LVEF, 67.0 ± 2.3%) and significantly attenuated LV dilatation (LV end-diastolic diameter, 4.0 ± 1.1 mm vs. cont-SkM, 4.5 ± 1.2 mm vs. MI-only, 4.8 ± 1.8 mm); this was associated with greater graft numbers, as shown by PCR for male-specificsmcygene. Enzyme zymography showed attenuated matrix metalloproteinase-2 and -9 up-regulation post-MI by either donor SkM type, although infarct-remote zone collagen was reduced only with sIL-1ra-SkM. These results suggest that SkM implantation improves cardiac function post-MI by modulation of adverse remodeling, and that this effect can be significantly enhanced by targeting IL-1 as a key upstream regulator of both adverse remodeling and graft cell death.
Cellular targets and mechanisms of nitros(yl)ation: An insight into their nature and kinetics
in vivo
Authors: Bryan Nathan S. , Rassaf Tienush , Maloney Ronald E. , Rodriguez Cynthia M. , Saijo Fumito , Rodriguez Juan R. , Feelisch Martin
Published: March 2004

There is mounting evidence that the established paradigm of nitric oxide (NO) biochemistry, from formation through NO synthases, over interaction with soluble guanylyl cyclase, to eventual disposal as nitrite/nitrate, represents only part of a richer chemistry through which NO elicits biological signaling. Additional pathways have been suggested that include interaction of NO-derived metabolites with thiols and metals to form
S
-nitrosothiols (RSNOs) and metal nitrosyls. Despite the overwhelming attention paid in this regard to RSNOs, little is known about the stability of these species, their significance outside the circulation, and whether other nitros(yl)ation products are of equal importance. We here show that N-nitrosation and heme-nitrosylation are indeed as ubiquitous as S-nitrosation
in vivo
and that the products of these reactions are constitutively present throughout the organ system. Our study further reveals that all NO-derived products are highly dynamic, have fairly short lifetimes, and are linked to tissue oxygenation and redox state. Experimental evidence further suggests that nitroso formation occurs substantially by means of oxidative nitrosylation rather than NO autoxidation, explaining why S-nitrosation can compete effectively with nitrosylation. Moreover, tissue nitrite can serve as a significant extravascular pool of NO during brief periods of hypoxia, and tissue nitrate/nitrite ratios can serve as indicators of the balance between local oxidative and nitrosative stress. These findings vastly expand our understanding of the fate of NO
in vivo
and provide a framework for further exploration of the significance of nitrosative events in redox sensing and signaling. The findings also raise the intriguing possibility that N-nitrosation is directly involved in the modulation of protein function.
Order in dense hydrogen at low temperatures
Authors: Edwards B. , Ashcroft N. W.
Published: March 2004

By increase in density, impelled by pressure, the electronic energy bands in dense hydrogen attain significant widths. Nevertheless, arguments can be advanced suggesting that a physically consistent description of the general consequences of this electronic structure can still be constructed from interacting but state-dependent multipoles. These reflect, in fact self-consistently, a disorder-induced localization of electron states partially manifesting the effects of proton dynamics; they retain very considerable spatial inhomogeneity (as they certainly do in the molecular limit). This description, which is valid provided that an overall energy gap has not closed, leads at a mean-field level to the expected quadrupolar coupling, but also for certain structures to the eventual emergence of dipolar terms and their coupling when a state of broken charge symmetry is developed. A simple Hamiltonian incorporating these basic features then leads to a high-density, low-temperature phase diagram that appears to be in substantial agreement with experiment. In particular, it accounts for the fact that whereas the phase I–II phase boundary has a significant isotope dependence, the phase II–III boundary has very little.
M-phase kinases induce phospho-dependent ubiquitination of somatic Wee1 by SCF
β-TrCP
Authors: Watanabe Nobumoto , Arai Harumi , Nishihara Yoshifumi , Taniguchi Makoto , Watanabe Naoko , Hunter Tony , Osada Hiroyuki
Published: March 2004

Wee1, the Cdc2 inhibitory kinase, needs to be down-regulated at the onset of mitosis to ensure rapid activation of Cdc2. Previously, we have shown that human somatic Wee1 (Wee1A) is down-regulated both by protein phosphorylation and degradation, but the underlying mechanisms had not been elucidated. In the present study, we have identified the β-transducin repeat-containing protein 1/2 (β-TrCP1/2) F-box protein-containing SKP1/Cul1/F-box protein (SCF) complex (SCF
β-TrCP1/2
) as an E3 ubiquitin ligase for Wee1A ubiquitination. Although Wee1A lacks a consensus DS(p)GXXS(p) phospho-dependent binding motif for β-TrCP, recognition of Wee1A by β-TrCP depended on phosphorylation, and two serine residues in Wee1A, S53 and S123, were found to be the most important phosphorylation sites for β-TrCP recognition. We have found also that the major M-phase kinases polo-like kinase 1 (Plk1) and Cdc2 are responsible for the phosphorylation of S53 and S123, respectively, and that in each case phosphorylation generates an unconventional phospho-degron (signal for degradation) that can be recognized by β-TrCP. Phosphorylation of Wee1A by these kinases cooperatively stimulated the recognition and ubiquitination of Wee1A by SCF
β-TrCP1/2
in vitro
. Mutation of these residues or depletion of β-TrCP by small-interfering RNA treatment increased the stability of Wee1A in HeLa cells. Moreover, our analysis indicates that β-TrCP-dependent degradation of Wee1A is important for the normal onset of M-phase
in vivo
. These results also establish the existence of a feedback loop between Cdc2 and Wee1A in somatic cells that depends on ubiquitination and protein degradation and ensures the rapid activation of Cdc2 when cells are ready to divide.
The AHNAKs are a class of giant propeller-like proteins that associate with calcium channel proteins of cardiomyocytes and other cells
Authors: Komuro Akihiko , Masuda Yutaka , Kobayashi Koichi , Babbitt Roger , Gunel Murat , Flavell Richard A. , Marchesi Vincent T.
Published: March 2004

To explore the function of the giant AHNAK molecule, first described in 1992 [Shtivelman, E., Cohen, F. E. & Bishop, J. M. (1992)
Proc. Natl. Acad. Sci. USA
89, 5472–5476], we created AHNAK null mice by homologous recombination. Homozygous knockouts showed no obvious phenotype, but revealed instead a second AHNAK-like molecule, provisionally designated AHNAK2. Like the original AHNAK, AHNAK2 is a 600-kDa protein composed of a large number of highly conserved repeat segments. Structural predictions suggest that the repeat segments of both AHNAKs may have as their basic framework a series of linked, antiparallel β-strands similar to those found in β-propeller proteins. Both AHNAKs appear to localize to Z-band regions of mouse cardiomyocytes and cosediment with membrane vesicles containing the dihydropyridine receptor, which is consistent with earlier reports that the AHNAKs are linked to L-type calcium channels and can be phosphorylated by protein kinase A. The localization of the AHNAKs in close proximity to transverse tubule membranes and Z-band regions of cardiac sarcomeres raise the possibility that they might be involved in regulating excitation/contraction coupling of cardiomyocytes, but other studies indicate that the association of AHNAKs with calcium channel proteins is more widespread. AHNAK2 is predicted to have a PDZ domain within its N-terminal, nonrepeating domain, which may mediate these interactions.
Disruption of Doppel prevents neurodegeneration in mice with extensive
Prnp
deletions
Authors: Genoud Nicolas , Behrens Axel , Miele Gino , Robay Dimitri , Heppner Frank L. , Freigang Stefan , Aguzzi Adriano
Published: March 2004

The
Prnp
gene encodes the cellular prion protein PrP
C
. Removal of its ORF does not result in pathological phenotypes, but deletions extending into the upstream intron result in cerebellar degeneration, possibly because of ectopic cis-activation of the
Prnd
locus that encodes the PrP
C
homologue Doppel (Dpl). To test this hypothesis, we removed
Prnd
from
Prnp
o/o
mice by transallelic meiotic recombination. Balanced
loxP
-mediated ablation yielded mice lacking both PrP
C
and Dpl (
Prn
o/o
), which developed normally and showed unimpaired immune functions but suffered from male infertility. However, removal of the
Prnd
locus abolished cerebellar degeneration, proving that this phenotype is caused by Dpl upregulation. The absence of compound pathological phenotypes in
Prn
o/o
mice suggests the existence of alternative compensatory mechanisms. Alternatively, Dpl and PrP
C
may exert distinct functions despite having partly overlapping expression profiles.
Crystal structure of RlmA
I
: Implications for understanding the 23S rRNA G745/G748-methylation at the macrolide antibiotic-binding site
Authors: Das Kalyan , Acton Thomas , Chiang Yiwen , Shih Lydia , Arnold Eddy , Montelione Gaetano T.
Published: March 2004

The RlmA class of enzymes (RlmA
I
and RlmA
II
) catalyzes N1-methylation of a guanine base (G745 in Gram-negative and G748 in Gram-positive bacteria) of hairpin 35 of 23S rRNA. We have determined the crystal structure of
Escherichia coli
RlmA
I
at 2.8-Å resolution, providing 3D structure information for the RlmA class of RNA methyltransferases. The dimeric protein structure exhibits features that provide new insights into its molecular function. Each RlmA
I
molecule has a Zn-binding domain, responsible for specific recognition and binding of its rRNA substrate, and a methyltransferase domain. The asymmetric RlmA
I
dimer observed in the crystal structure has a well defined W-shaped RNA-binding cleft. Two
S
-adenosyl-
l
-methionine substrate molecules are located at the two valleys of the W-shaped RNA-binding cleft. The unique shape of the RNA-binding cleft, different from that of known RNA-binding proteins, is highly specific and structurally complements the 3D structure of hairpin 35 of bacterial 23S rRNA. Apart from the hairpin 35, parts of hairpins 33 and 34 also interact with the RlmA
I
dimer.
Transcriptional switch by activating transcription factor 2-derived peptide sensitizes melanoma cells to apoptosis and inhibits their tumorigenicity
Authors: Bhoumik Anindita , Jones Nic , Ronai Ze'ev
Published: March 2004

The notorious resistance of melanoma cells to drug treatment can be overcome by expression of a 50-aa peptide derived from activating transcription factor 2 (ATF2
50–100
). Here we demonstrate that ATF2
50–100
induced apoptosis by sequestering ATF2 to the cytoplasm, thereby inhibiting its transcriptional activities. Furthermore, ATF2
50–100
binds to c-Jun N-terminal kinase (JNK) and increases its activity. Mutation within ATF2
50–100
that impairs association with JNK and the inhibition of JNK or c-Jun expression by RNA interference (RNAi) reduces the degree of ATF2
50–100
-induced apoptosis. In contrast, TAM67, a dominant negative of the Jun family of transcription factors, or JunD RNAi attenuates sensitization of melanoma cells expressing ATF2
50–100
to apoptosis after treatment with anisomycin, which is used as a model drug. Mutations within the JNK binding region of ATF2
50–100
or expression of TAM67 or JunD RNAi attenuates inhibition of melanoma's tumorigenicity by ATF2
50–100
. We conclude that inhibition of ATF2 in concert with increased JNK/Jun and JunD activities is central for the sensitization of melanoma cells to apoptosis and inhibition of their tumorigenicity.
Membrane-mediated structural transitions at the cytoplasmic face during integrin activation
Authors: Vinogradova Olga , Vaynberg Julia , Kong Xiangming , Haas Thomas A. , Plow Edward F. , Qin Jun
Published: March 2004

Cytoplasmic face-mediated integrin inside-out activation remains a paradigm in transmembrane signal transduction. Emerging evidence suggests that this process involves dissociation of the complex between the integrin cytoplasmic tails; however, a dynamic image of how it occurs on the membrane surface remains elusive. We show here that, whereas membrane-proximal helices of integrin α/β cytoplasmic tails associate in cytoplasm-like aqueous medium, they become partially embedded into membranemimetic micelles when unclasped. Membrane embedding induces substantial structural changes of the cytoplasmic tails as compared to their aqueous conformations and suggests there may be an upward movement of the membrane-proximal helices into the membrane during their separation. We further demonstrate that the β3 tail exhibits additional membrane binding site at its C terminus containing the NPLY motif. Talin, a key intracellular integrin activator, recognizes this site as well as the membrane-proximal helix, thereby promoting cytoplasmic tail separation along the membrane surface. These data provide a structural basis of membrane-mediated changes at the cytoplasmic face in regulating integrin activation and signaling.
A global suppressor motif for p53 cancer mutants
Authors: Baroni Timothy E. , Wang Ting , Qian Hua , Dearth Lawrence R. , Truong Lan N. , Zeng Jue , Denes Alec E. , Chen Stephanie W. , Brachmann Rainer K.
Published: March 2004

The transcription factor and tumor suppressor protein p53 is frequently inactivated in human cancers. In many cases,
p53
gene mutations result in high levels of inactive, full-length p53 protein with one amino acid change in the core domain that recognizes p53 DNA-binding sites. The ability to endow function to mutated p53 proteins would dramatically improve cancer therapy, because it would reactivate a central apoptotic pathway. By using genetic strategies and p53 assays in yeast and mammalian cells, we identified a global suppressor motif involving codons 235, 239, and 240. These intragenic suppressor mutations, either alone or in combination, restored function to 16 of 30 of the most common p53 cancer mutants tested. The 235-239-240 suppressor motif establishes that manipulation of a small region of the core domain is sufficient to activate a large number of p53 cancer mutants. Understanding the structural basis of the rescue mechanism will allow the pursuit of small compounds able to achieve a similar stabilization of p53 cancer mutants.
Evolving potassium channels by means of yeast selection reveals structural elements important for selectivity
Authors: Bichet Delphine , Lin Yu-Fung , Ibarra Christian A. , Huang Cindy Shen , Yi B. Alexander , Jan Yuh Nung , Jan Lily Yeh
Published: March 2004

Potassium channels are widely distributed. To serve their physiological functions, such as neuronal signaling, control of insulin release, and regulation of heart rate and blood flow, it is essential that K
+
channels allow K
+
but not the smaller and more abundant Na
+
ions to go through. The narrowest part of the channel pore, the selectivity filter formed by backbone carbonyls of the GYG-containing K
+
channel signature sequence, approximates the hydration shell of K
+
ions. However, the K
+
channel signature sequence is not sufficient for K
+
selectivity. To identify structural elements important for K
+
selectivity, we randomly mutagenized the G protein-coupled inwardly rectifying potassium channel 3.2 (GIRK2) bearing the S177W mutation on the second transmembrane segment. This mutation confers constitutive channel activity but abolishes K
+
selectivity and hence the channel's ability to complement the K
+
transport deficiency of Δ
trk1
Δ
trk2
mutant yeast. S177W-containing GIRK2 mutants that support yeast growth in low-K
+
medium contain multiple suppressors, each partially restoring K
+
selectivity to S177W-containing double mutants. These suppressors include mutations in the first transmembrane segment and the pore helix, likely exerting long-range actions to restore K
+
selectivity, as well as a mutation of a second transmembrane segment residue facing the cytoplasmic half of the pore, below the selectivity filter. Some of these suppressors also affected channel gating (channel open time and opening frequency determined in single-channel analyses), revealing intriguing interplay between ion permeation and channel gating.
Inhibition of Adipogenesis by Ghrelin
Authors: Zhang Weizhen , Zhao Lili , Lin Theodore R. , Chai Biaoxin , Fan Yongyi , Gantz Ira , Mulholland Michael W.
Published: March 2004

Ghrelin, a novel gastric hormone, regulates food intake and energy metabolism via central mechanisms. The peripheral effect of ghrelin on adiposity is poorly understood. We established a stable 3T3-L1 cell line expressing ghrelin to study the direct effect of ghrelin on adipogenesis. Cells overexpressing ghrelin demonstrate significantly attenuated differentiation of preadipocytes into adipocytes. Expression of peroxisome proliferator-activator receptor-γ is significantly inhibited as demonstrated by decrease of peroxisome proliferator-activator receptor-γ mRNA and protein. Both ghrelin overexpression and exogenous ghrelin stimulate cell proliferation. Phosphorylation of mitogen-activated protein kinase is increased after treatment of cells with ghrelin. Ghrelin binding activity is demonstrated in both native and ghrelin-overexpressing 3T3-L1 cells by radiolabeled ghrelin, although reverse transcription-polymerase chain reaction with the primer sequence of the previously identified ghrelin receptor subtypes detected no signal. Our results demonstrate that ghrelin inhibits adipogenesis by stimulation of cell proliferation via the mediation of a ghrelin receptor, likely a novel unidentified subtype.
The Palmitoyltransferase of the Cation-dependent Mannose 6-Phosphate Receptor Cycles between the Plasma Membrane and Endosomes
Authors: Stöckli Jacqueline , Rohrer Jack
Published: March 2004

The cation-dependent mannose 6-phosphate receptor (CD-MPR) mediates the transport of lysosomal enzymes from the trans-Golgi network to endosomes. Evasion of lysosomal degradation of the CD-MPR requires reversible palmitoylation of a cysteine residue in its cytoplasmic tail. Because palmitoylation is reversible and essential for correct trafficking, it presents a potential regulatory mechanism for the sorting signals within the cytoplasmic domain of the CD-MPR. Characterization of the palmitoylation performing an in vitro palmitoylation assay by using purified full-length CD-MPR revealed that palmitoylation of the CD-MPR occurs enzymatically by a membrane-bound palmitoyltransferase. In addition, analysis of the localization revealed that the palmitoyltransferase cycles between endosomes and the plasma membrane. This was identified by testing fractions from HeLa cell homogenate separated on a density gradient in the in vitro palmitoylation assay and further confirmed by in vivo labeling experiments by using different treatments to block specific protein trafficking steps within the cell. We identified a novel palmitoyltransferase activity in the endocytic pathway responsible for palmitoylation of the CD-MPR. The localization of the palmitoyltransferase not only fulfills the requirement of our hypothesis to be a regulator of the intracellular trafficking of the CD-MPR but also may affect the sorting/activity of other receptors cycling through endosomes.
Non-linear conversion between genetic and physical chromosomal distances
Authors: Voigt Claudia , Möller Steffen , Ibrahim Saleh M. , Serrano-Fernández Pablo
Published: March 2004

Summary: A supervised nonlinear interpolation significantly improves the reliability of conversions from genetic distances to physical distances as compared with the linear ones. A webaccessible application was created that addresses this question with a graphical presentation that may be wrapped by local installations.

Motivation: Genetic linkage maps and radiation hybrid (RH) maps are based on the rate of uncoupling between linked genetic markers. These are usually measured in centiMorgan (cM) when uncoupling is originated by natural recombination or in centiRay (cR) for chromosomes that are irradiated artificially to separate the markers. Physical maps arise from genome-wide DNA sequencing and are measured in bp.

This work was originally motivated as an extension of the software application Expressionview (Fischer et al., 2003), exploring its spectrum of appliance combining different mapping systems. The relationship between physical and genetic maps is known to be not always linear (Yu et al., 2001). The shift from the linear model seems to depend on local idiosyncrasies of the chromosomes and the kind of genetic map used. The present application addresses this problem for the first time.

Availability:  http://qtl.pzr.uni-rostock.de/cartographer.php