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L Weiner

Publications and source records attributed to L Weiner.

At least 55 records · Page 3Linked to original sources
PubMed1994

Two-state transition between molten globule and unfolded states of acetylcholinesterase as monitored by electron paramagnetic resonance spectroscopy.

Cys-231 of Torpedo californica acetylcholinesterase (EC 3.1.1.7) was selectively labeled with the mercury derivative of a stable nitroxyl radical. In 1.5 M guanidinium chloride, this conjugate exists in a molten globule state (MG), whereas in 5 M denaturant, it is in an unfolded state (U). The transition between the two states is reversible. In the MG, the label is highly immobilized, whereas in the U, it is almost freely rotating. The clearly distinct electron paramagnetic resonance (EPR) spectra of the two states permits the study of this transition. Upon elevating the guanidinium chloride concentration, a decrease in the EPR signal of the MG occurs concomitantly with an increase in the U signal, the total intensity of the EPR spectra remaining constant. This behavior is characteristic of a two-state transition. The thermodynamic characteristics of this transition (delta G0 and m), whether estimated directly from the EPR data or from both CD and fluorescence data analyzed by assuming a two-state scheme, are in good agreement.

Acetylcholinesterase↗
PubMed1994

A "molten globule" of Torpedo acetylcholinesterase undergoes thiol-disulfide exchange.

Torpedo acetylcholinesterase is a disulfide-linked homodimer containing three intramolecular disulfide bonds, as well as a single free thiol on Cys-231. We report that in a "molten globule" state, produced by 1.5 M guanidine hydrochloride, this enzyme undergoes rapid intramolecular thiol-disulfide exchange, in the absence of reducing agents, resulting in the production of novel species. Most strikingly, this results in appearance of enzyme monomers. Chemical modification of the free thiol group prevents these changes. Unfolded acetylcholinesterase, namely in 5 M guanidine hydrochloride, also undergoes intramolecular thiol-disulfide exchange, including production of enzyme monomers, but at a much lower rate. Our data show that the molten globule state, in contrast to the native and unfolded states, is both compact and flexible, thus being especially amenable to thiol-disulfide exchange.

Acetylcholinesterase↗
PubMed1994

Preoperative chemoradiation for adenocarcinoma of the pancreas and duodenum.

PURPOSE: This study was designed to evaluate the effects of preoperative chemoradiation on resectability, response, local control, and survival in patients with local or local-regional involvement from carcinoma of the pancreas or cancer of the duodenum and to assess the associated toxicity of such treatment. METHODS AND MATERIALS: This prospective pilot study of preoperative chemoradiation was initiated in 1986 for patients with clinical evidence of adenocarcinoma of the pancreas or duodenum without evidence of distant metastases. Radiation was given at 1.8 Gy per day to a total dose of 50.4 Gy. Two cycles of chemotherapy were given concurrent with radiation. On days 2-5 and 29-32, 5-fluorouracil (1 gm/m2/24 h x 4 days) was given, while mitomycin-C (10 mg/m2) was given on day 2 only. Surgical resection was 4-6 weeks following completion of chemoradiation. Thirty-one patients (17 male and 14 female) were entered on the protocol with a median potential follow-up of 4.5 years (range 6 months to 7.5 years). The median age was 64 years (range 32-73 years). Twenty-seven patients had pancreatic cancer (25 head, two body), while four patients had carcinoma arising from the duodenum. Twenty-one patients were initially judged to be unresectable and ten potentially resectable prior to chemoradiation. RESULTS: Twenty-nine of 31 patients completed the entire course of radiation and both cycles of chemotherapy. Acute toxicity from chemoradiation consisted of nausea, vomiting, diarrhea, stomatitis, or hematologic suppression which was moderate to severe (Grade 3 or 4) in seven patients (23%). One patient died of sepsis following the first week of therapy. Seventeen patients (55%) underwent curative resection with subtotal or total pancreatectomy or Whipple resection (four duodenum, 13 pancreas) and two (2/17) had pathologic nodal involvement, while (0/17) none had involved margins. A complete pathologic response was seen in all four (4/4) patients with duodenal cancer and in none (0/13) with pancreatic cancer who underwent resection. The median postoperative hospitalization stay was 22 days (range 4-144 days). Of 17 patients who underwent curative resection, there were two postoperative mortalities (12%). Late complications have included abscess, one; and nonmalignant ascites, five. Ten of the 31 patients are alive. For patients with pancreatic cancer the median survival is 9 months, while survival at 1 year and 3 years are 36% and 19% overall and 60% and 43% at 1 and 3 years for those undergoing resection. Six of the 27 patients (22%) with pancreatic cancer are alive without recurrence. All four patients with duodenal cancer are alive without recurrence (12 months, 23 months, 35 months, 90 months). CONCLUSION: Preoperative chemoradiation for cancer of the pancreas and duodenal region was relatively well-tolerated and enhanced resectability and downstaging of nodal metastases were suggested. The 3-year survival, particularly in patients who underwent resection, was high. For these reasons the applicability of this treatment regimen for pancreatic cancer is presently being studied in a group-wide multi-institutional Phase II trial. Chemoradiation for duodenal cancer has produced a complete pathologic response in all patients and survival has been excellent, suggesting efficacy of this regimen for duodenal cancer.

Adenocarcinoma↗
PubMed1994

Role of an Escherichia coli stress-response operon in stationary-phase survival.

The phage shock protein operon (pspABCE) of Escherichia coli is strongly expressed in response to stressful environmental conditions, such as heat shock, ethanol treatment, osmotic shock, and filamentous phage infection. We show that bacteria lacking the pspABC genes exhibit a substantial decrease in the ability to survive prolonged incubation in stationary phase under alkaline conditions (pH 9). The psp mutant bacteria grow approximately as well as wild-type strains in the alkaline medium, and stationary-phase survival of the psp mutants improves substantially at pH values closer to the optimal growth range (pH 6-8). In late stationary-phase (1- to 2-day-old) cells, the operon can be strongly induced under certain conditions, and PspA can become one of the most highly expressed bacterial proteins. The combination of stationary-phase starvation and alkaline pH is likely to place a severe strain on the maintenance of endogenous energy sources, and, consistent with these effects, we find that psp expression is also induced by uncouplers of oxidative phosphorylation and other agents that interfere with energy production. The death rate of psp mutants in stationary phase is accelerated by the presence of wild-type bacteria in the same culture, suggesting that the psp operon may play a significant role in enabling E. coli to compete for survival under nutrient- or energy-limited conditions.

Bacterial Proteins↗
PubMed1994

Oxidative stress transforms acetylcholinesterase to a molten-globule-like state.

Exposure of purified acetylcholinesterase from Torpedo california to a system generating oxygen radicals (viz. ascorbic acid/Fe(EDTA)2/H2O2) inactivated the enzyme. The enzyme retained its native dimeric form, but electrophoresis under denaturing conditions showed some cleavage of peptide bonds. Spectroscopic characterization revealed a shift to the red in the intrinsic fluorescence emission peak, a large decrease in molar ellipticity in the near UV with a much smaller decrease in the far UV, and increased binding of the amphiphilic probe, 1-anilino-8-naphthalene sulfonate, all relative to native enzyme. The treated enzyme was also highly susceptible to proteolysis. These data show that oxygen radical treatment converts acetylcholinesterase to a partially unfolded state, which retains most of its secondary structure but lacks substantial tertiary structure, thus resembling a 'molten globule' state. This model system may offer a mechanism for explaining the consequences of oxidative stress in vivo: partially unfolded proteins generated by oxidative stress may interact with molecular chaperons of the heat shock family, thus activating the heat-shock factor and, thereby, activating heat-shock genes.

Acetylcholinesterase↗
PubMed1993

Electron leakage from the mitochondrial NADPH-adrenodoxin reductase-adrenodoxin-P450scc (cholesterol side chain cleavage) system.

In electron (e-) transfer systems some e- may "leak," reducing O2 to a superoxide radical. This study examined the sites and kinetics of e-leakage from the mitochondrial P450scc system. Adrenodoxin reductase alone oxidized NADPH, reducing O2 to a superoxide radical at a very low rate. However, the reductase-adrenodoxin system reduced O2 at a rapid steady-state rate with Michaelis-Menten dependence on [adrenodoxin](Vmax = 3.5 micro M e-/min). After depletion of NADPH, reduced adrenodoxin was oxidized (autooxidation) with pseudo first order kinetics and the rate of e- transfer decreased 10-fold. Ca2+ (< 1 mM) stimulated e- leakage in both phases. The reductase-adrenodoxin-P450scc system exhibited the highest rate of leakage (Vmax = 7.8 microM e-/min). At low [adrenodoxin] the majority of e-leaked through P450scc and not through adrenodoxin. In the presence of the substrate, cholesterol, leakage drastically decreased to <0.5 microM e-/min. These results indicate that the mitochondrial P450 systems can leak e-, producing O2 derived free radicals. Reduction of leakage during P450scc conversion of cholesterol to pregnenolone provides a clue to understanding physiological mechanisms that control e-leakage. These may include coregulation of NADPH and cholesterol availability to the P450scc system and a system of antioxidants for quenching the oxygen radicals.

Adrenal Cortex↗
PubMed1993

Effects of small doses of bisoprolol on blood pressure and lipoprotein concentrations in hypertensive patients.

The effects of bisoprolol 2.5 and 5 mg per day on blood pressure, and lipoprotein and apolipoprotein concentrations were compared in 18 newly detected hypertensives in a double-blind, crossover study. All treatment results were related to the values at the end of a four-week placebo run-in period. Each of the two following treatment periods lasted for 3 months. The systolic and diastolic pressures in the supine position were reduced by 19.5/11.7 mm Hg and 14.6/10.4 mm Hg by 2.5 and 5 mg bisoprolol per day, respectively, with no significant difference in effect. Supine heart rate was reduced by 4.7 and 8.2 beats.min-1, respectively, (P = 0.0517 for different effects). The cholesterol concentration in low-density (LDL) and high-density (HDL) lipoproteins was reduced during both regimens, by about 0.3 and 0.1 mmol.l-1, respectively, difference not significant. Triglyceride concentrations were not significantly affected during either regimen. We conclude that, in this study population, treatment with bisoprolol 2.5 mg per day was equally effective as 5.0 mg per day in reducing blood pressure. The effects on lipoprotein concentrations were small and included an unexpected reduction in LDL-cholesterol concentration. A low dose of a highly selective beta-adrenoceptor blocker like bisoprolol appears to retain the blood pressure reducing capacity and has lost most of the unfavourable effects on lipoproteins characteristic of higher doses.

Adult↗
PubMed1993

Pilon fractures: treatment with combined internal and external fixation.

The purpose of this study was to prospectively evaluate the use of limited internal fixation and the application of a hybrid external fixator (tensioned wires distally and 5.0 mm half pins proximally attached to a semicircular frame without crossing the ankle joint) in the treatment of severe distal tibia fractures. This technique involves accurate reduction and fixation of the intraarticular component through an incision based over a fracture site followed by stabilization of the metaphysis with the hybrid external fixator. We studied 26 patients 15-55 years of age who were followed for 8-36 months. All fractures were within 5 cm of the joint. Seventeen fractures were intraarticular, nine extraarticular, and six open. Eleven patients required bone grafting. The average time to healing was 4.2 months. Using clinically based criteria, there were 81% good and excellent results overall, 70.5% for the 17 intraarticular fractures, and 69% for Ruedi type III fractures. Complications included one superficial and one deep infection, one 10 degrees varus malunion, and three pin tract infections. This method yielded results comparable with previous studies while reducing the amount of soft tissue dissection necessary for the placement of large plates. Soft tissue complications were infrequent and the goals of early motion and fracture stability were not sacrificed.

Adolescent↗
PubMed1992

The application of pH-sensitive spin labels to studies of surface potential and polarity of phospholipid membranes and proteins.

The effects of pH titration on the EPR spectra of imidazolidine nitroxides located at the surface of mixed bilayers composed of dimyristoylphosphatidylglycerol (DMPG) and dimyristoylphosphatidylcholine (DMPC), and at the surface of the protein, human serum albumin (HSA), have been investigated. It is found that the shift in pKa of the amino group of the imidazolidine radical from its value of 4.6 in water depends both on the interfacial polarity (delta pKapol) and on the electrostatic surface potential (delta pKael) when it is positioned at the bilayer/water interface by an anchoring hydrocarbon tail. The polarity shift is determined to be: delta pKapol = -1.3 units at the surface of DMPC bilayers at 17 degrees C, corresponding to an effective interfacial dielectric constant of epsilon approximately 37, and depends on the temperature with a coefficient of d delta pKapol/dT approximately -0.01 per degree. The electrostatic shift at the surface of DMPG bilayers is delta pKael = +1.6 units in 0.1 M KCl, which corresponds to an electrostatic surface potential of -95 mV. This electrostatic shift depends strongly both on ionic strength and on the fraction of charged lipid in the DMPC/DMPG mixtures, in a manner that agrees with the predictions of electrostatic double-layer theory. It is found that the shift in pKa of an imidazolidine radical covalently bound at the surface of HSA is determined mainly by the surface electrostatics (delta pKapol approximately 0) and corresponds to an electrostatic potential of +33 mV in 0.01 M KCl at a pH below the isoelectric point of the protein.

Dimyristoylphosphatidylcholine↗
PubMed1992

Pediatricians' perspectives on fetal alcohol syndrome.

Since the identification of fetal alcohol syndrome (FAS) in 1973, significant inroads have been made towards understanding the effects of alcohol on fetal development. However, it is not clear if these findings are considered clinically relevant by pediatricians. This survey was designed to assess clinical knowledge, practice, and attitudes concerning alcohol-related birth defects. Data were collected in a questionnaire that was mailed to 234 randomly selected Massachusetts pediatricians. Responses suggest that a substantial proportion of pediatricians have knowledge about the effects of alcohol on pregnancy. However, many considered themselves unprepared to deal with this topic. More physicians suspected FAS/FAE than made the diagnosis. Almost three fourths reported they would find professional education in this area helpful. Broader dissemination of research findings in clinically relevant formats and improving the sense of preparedness among pediatricians have the potential to improve the care of children born to heavily drinking pregnant women.

Attitude of Health Personnel↗
PubMed1991

Characterization and sequence of the Escherichia coli stress-induced psp operon.

We describe a new Escherichia coli operon, the phage shock protein (psp) operon, which is induced in response to heat, ethanol, osmotic shock and infection by filamentous bacteriophages. The operon includes at least four genes: pspA, B, C and E. PspA associates with the inner membrane and has the heptad repeats characteristic of proteins that can form coiled coils. The operon encodes a factor that activates psp expression, and deletion analyses indicate that this protein is PspC; PspC is predicted to possess a leucine zipper, a motif present in many eukaryotic transcription factors. The pspE gene is expressed in response to stress as part of the operon, but is also transcribed from its own promoter under normal conditions. In vitro studies suggest that PspA and C are modified in vivo. Expression of the psp genes does not require the heat shock sigma factor, sigma32. The increased duration of psp induction in a sigma32 mutant suggests that a product (or products) of the heat shock response down-regulates expression of the operon.

Amino Acid Sequence↗
PubMed1991

Antierythrocyte autoantibody formation after therapy with interleukin-2 and gamma-interferon.

The cardiovascular, renal, pulmonary, and dermatologic toxicities of interleukin-2 (IL-2) and gamma-interferon (IFN) are well described. However, autoimmune toxicities have only recently been noticed. The authors report the development of warm autoantibodies against erythrocytes in a patient receiving IL-2 (3.75 x 10(6) cetus units/m2 intravenous bolus three times per week) and gamma-IFN (0.1 mg/m2 subcutaneously three times per week) for metastatic renal cell carcinoma. Other potential causes of autoantibody formation, such as drugs, infection, and collagen vascular disease, were excluded. Both gamma-IFN and IL-2 have the potential to trigger or exacerbate autoimmunity due to either aberrant expression of restricted antigens or inhibition of normal cellular immune suppressor mechanisms.

Autoantibodies↗
PubMed1991

Stress-induced expression of the Escherichia coli phage shock protein operon is dependent on sigma 54 and modulated by positive and negative feedback mechanisms.

The phage shock protein (psp) operon of Escherichia coli is strongly induced in response to heat, ethanol, osmotic shock, and infection by filamentous bacteriophages. The operon contains at least four genes--pspA, pspB, pspC, and pspE--and is regulated at the transcriptional level. We report here that psp expression is controlled by a network of positive and negative regulatory factors and that transcription in response to all inducing agents is directed by the sigma-factor sigma 54. Negative regulation is mediated by both PspA and the sigma 32-dependent heat shock proteins. The PspB and PspC proteins cooperatively activate expression, possibly by antagonizing the PspA-controlled repression. The strength of this activation is determined primarily by the concentration of PspC, whereas PspB enhances but is not absolutely essential for PspC-dependent expression. PspC is predicted to contain a leucine zipper, a motif responsible for the dimerization of many eukaryotic transcriptional activators. PspB and PspC, though not necessary for psp expression during heat shock, are required for the strong psp response to phage infection, osmotic shock, and ethanol treatment. The psp operon thus represents a third category of transcriptional control mechanisms, in addition to the sigma 32- and sigma E-dependent systems, for genes induced by heat and other stresses.

Bacterial Proteins↗
PubMed1991

Effects of early tangential excision and grafting on survival after burn injury.

This report quantifies the increase of burn survival, which we believe is associated with the use of early tangential excision and grafting as opposed to conventional therapy in adult patients with burns. To quantify the increase, we compared the lethal area 50 of various age groups with that of previous historical studies that used other techniques. These data suggest that early excision and grafting are associated with a marked improvement in survival in patients 41 to 60 years of age with 20 to 65 per cent total body surface area burns.

Adult↗