Search PubMed⌕ Search

Biomedical subjects

M Gutman

Publications and source records attributed to M Gutman.

At least 55 records · Page 3Linked to original sources
PubMed1997

Mechanism of proton entry into the cytoplasmic section of the proton-conducting channel of bacteriorhodopsin.

Bacteriorhodopsin is the light-driven proton-pumping protein of Halobacterium salinarum that extracts protons from the well-buffered cytoplasmic space within the time limits set by the photocycle turnover. The specific mechanism of the proton uptake by the cytoplasmic surface of the protein was investigated in this study by the laser-induced proton pulse technique. The purple membrane preparations were labeled by fluorescein at two residues (36 or 38) of the cytoplasmic surface of the protein, sites that are close to the orifice of the proton-conducting channel. The membranes were pulsed by protons discharged from photoexcited pyranine [Nachliel, E., Gutman, M., Kiryati, S., and Dencher, N.A. (1996) Proc. Nat Acad. Sci. U.S.A. 93, 10747-10752). The reaction of the discharged protons with the pyranine anion and the fluorescein was measured with sub-microsecond resolution. The experimental signals were reconstructed through numeric integration of differential rate equations which quantitated the rates of all proton transfer reactions between all reactants present in the system. The interaction of protons with the orifice of the cytoplasmic channel is enhanced by the exposed carboxylates of the protein. A cluster of three carboxylates acts as a strong proton attractor site while one carboxylate, identified as D36, acts as a mediator that delivers the proton to the channel. The combination of these reactions render the surface of the protein with properties of a proton-collecting antenna. The size of the collecting area is less than that of the protein's surface.

Bacteriorhodopsins↗
PubMed1997

Expression of MCAM/MUC18 by human melanoma cells leads to increased tumor growth and metastasis.

The cell surface adhesion molecule MCAM (MUC18) is strongly expressed by advanced primary and metastatic melanomas but is weaker and less frequent in nevus cells. Previous studies have shown that MCAM expression correlates with tumor thickness and metastatic potential of human melanoma cells in nude mice. To provide direct evidence that MCAM plays a role in tumor growth and metastasis of human melanoma, the nonmetastatic MCAM-negative primary cutaneous melanoma SB-2 cells were transfected with MCAM cDNA and analyzed subsequently for changes in their tumorigenic and metastatic potential. Enforced expression of MCAM in SB-2 cells rendered them highly tumorigenic and increased their metastatic potential in nude mice as compared with parental and control transfected cells. The transfected cells displayed increased homotypic adhesion, increased attachment to human endothelial cells, decreased ability to adhere to laminin, and increased invasiveness through Matrigel-coated filters. Anti-MCAM monoclonal antibody reversed these functions in the transfected cells but not in control cells. The above changes in function attributed to the expression of MCAM may underlie the contribution of MCAM/MUC18 to the malignant phenotype.

Animals↗
PubMed1997

High dose tumor necrosis factor-alpha and melphalan administered via isolated limb perfusion for advanced limb soft tissue sarcoma results in a >90% response rate and limb preservation.

BACKGROUND: Recombinant tumor necrosis factor-alpha (rTNF-alpha) is a highly potential antineoplastic agent. However, its systemic administration in humans has resulted in a life-threatening septic shock-like syndrome, and its use has been abandoned. The administration of high dose rTNF-alpha and melphalan via isolated limb perfusion (ILP) eliminated the systemic side effects and was shown to be very effective for metastatic melanoma confined to the limb. The purpose of the current study was to assess the role of rTNF-alpha and melphalan administered via ILP in patients with soft tissue sarcoma. Amputation is unavoidable in 10% of these patients despite aggressive conventional therapy. Limb preservation was the objective in this select group of candidates for amputation or mutilating surgery. METHODS: During a 36-month period, 35 patients with high grade soft tissue sarcoma underwent 41 ILPs with high dose rTNF-alpha (3-4 mg) and melphalan (1-1.5 mg/kg). There were 21 males and 14 females. The mean age was 48 years (range, 14-80 years). Histologic subtypes included malignant fibrous histiocytoma, synovial, liposarcoma, malignant schwannoma, desmoid, clear cell, epithelioid, rhabdomyosarcoma, leiomyosarcoma, and unclassifiable. Twenty-one patients presented with recurrent and 14 with very extensive primary tumors. The tumors were located in the upper extremity in 8 patients and in the lower extremity in 27 patients. Twenty-five patients were candidates for amputation and 10 for extensive mutilating surgery. ILP was performed via the corresponding vessels proximal to the tumor. Six patients with partial response (PR) insufficient to render them resectable underwent a second ILP. With the exception of 4 of 9 patients with multifocal lesions and 1 who refused surgery, resection of the residual tumor or tumor bed or limb was performed 6-8 weeks after ILP. RESULTS: Marked tumor softening occurred within 48 hours, and in tumors protruding through the skin hemorrhagic necrosis was evident within 24 hours. The overall response rate was 91%. Thirteen patients (37%) had a complete response and 19 (54%) had a PR; in 5 of these patients > 90% necrosis of the tumor occurred. In 3 patients (8.5%), only minimal regression was observed (stabilization of disease). The rate of limb sparing was 85% (29 of 34 patients). CONCLUSIONS: The combination of high dose rTNF-alpha and melphalan given via ILP appears to be effective in patients with advanced soft tissue sarcoma confined to the limb, achieving a high response rate and limb preservation.

Adolescent↗
PubMed1997

Time-resolved dynamics of proton transfer in proteinous systems.

The dynamics of proton dissociation from an acidic moiety and its subsequent dispersion in the bulk is regulated by the physical chemical properties of the solvent. The solvent has to provide a potential well to accommodate the discharged proton, screen it from the negative charge of the conjugated base, and provide an efficient mode for the diffusion of the proton to the bulk. On measuring the dynamics of proton dissociation in the time-resolved domain, the kinetic analysis of the reaction can quantitate the properties of the immediate environment. In this review we implement the kinetic analysis for evaluating the properties of small cavities in proteins and the diffusion of protons within narrow channels. On the basis of this analysis,we discuss how the clustering of proton-binding sites on a surface can endow the surface with enhanced capacity to attract protons and to funnel them toward a specific site.

Journal Article↗
PubMed1997

Gut decontamination reduces bowel ischemia-induced lung injury in rats.

OBJECTIVE: To evaluate the effects of gut decontamination on endotoxin, tumor necrosis factor (TNF) levels, and the associated lung injury in a rat model of bowel ischemia. SUMMARY BACKGROUND DATA: Gut ischemia induces disruption of the intestinal mucosal barrier, allowing translocation of bacteria and endotoxin into the blood, which may trigger a systemic inflammatory response and lung injury. METHODS: Thirty anesthetized rats were randomized into three groups: (1) ischemia-reperfusion (I/R) alone (a 60-min superior mesenteric artery occlusion and 4 h of reperfusion, n=10); (2) rats that underwent gut decontamination prior to ischemia (I/R+GD, n=10); and (3) control rats (sham operated, n=10). Serum levels of lipopolysaccharide (LPS) and TNF were measured at the end of the experiment. Lung permeability was measured using bovine serum albumin labeled with 125I, and organ injury was assessed histologically. RESULTS: One hour of bowel ischemia and 4 h of reperfusion (I/R) led to elevations of blood LPS and TNF levels of 0.33+/-0.005 EU/mL and 173+/-56 pg/mL, which were higher than the sham group (p<0.01). Gut decontamination (I/R+GD) significantly attenuated the LPS and TNF generation, 0.09+/-0.005 and 56.2+/-6 pg/mL (p<0.01). Lung injury as assessed by pulmonary permeability index was also reduced by gut decontamination, 2.1+/-0.42 vs 5.3+/-0.82 in the control group (p<0.03). Surprisingly no difference was detected in the number of pulmonary neutrophils in sequestration between the groups. CONCLUSIONS: Our data suggest that gut decontamination can reduce the generation of LPS, TNF, and the severity of lung damage that often follows ischemia and reperfusion of the intestine in rats.

Animals↗
PubMed1997

Lessons learned from a mentoring program for teenage mothers.

PURPOSE: This study evaluated a mentoring program designed to decrease the risk of repeat pregnancy among unmarried primiparous teens, ages 12-19. METHODS: Adolescents (n = 110) completed a battery that assessed sexual/contraceptive behavior; psychological adjustment; and attitudes towards school. Teens were then randomly assigned to a mentor or control group, and reassessed at 6, 12, 18, and 24 months. Mentored teens received social support and assistance dealing with community agencies from mentors who were trained community volunteers. RESULTS AND CONCLUSIONS: At baseline, mentor and control teens had similar sexual histories, school achievement profiles (percentage enrolled, cumulative grade point average), and anticipated being the same age when they had a second child. At 24 months (n = 81), most mentor and control teens were making progress in school. Fifty percent had graduated or had advanced two grades; 10 of the 16 graduating teens were seeking additional education. However, the mentoring program did not significantly impact repeat pregnancy rates. At 33 months, 66.0% of the mentored teens and 68.8% of the control teens had experienced a repeat pregnancy. Thirty-six percent of teens had one repeat pregnancy; 24% had two or more pregnancies. Sixty-two percent of the pregnancies with known resolution (89) resulted in live births; 26% were aborted. In providing this mentoring program, several important lessons were learned.

Adolescent↗
PubMed1997

Synergism of tumor necrosis factor-alpha and melphalan in systemic and regional administration: animal study.

Tumor necrosis factor (TNF) is a highly cytotoxic cytokine. However, due to its severe side effects, the only clinical situation allowing its administration in humans is isolated limb perfusion (ILP). Early studies have shown that TNF alone is of limited efficacy even at high doses via ILP, and that a chemotherapeutic agent needs to be added. The most commonly used drug in this setting is melphalan which is considered to be synergistic with TNF. However, since melphalan has not been commonly used in sarcoma, we believed that confirmation of its synergistic effect with TNF in an experimental sarcoma model could prove valuable for future drug choice. B16F10 melanoma and CT26 colon carcinoma cells were injected subcutaneously (s.c.) into mice, while GF fibrosarcoma cells were injected s.c. into the hindleg of Wistar rats. The animals were then divided into four treatment groups: TNF alone, melphalan alone, TNF and melphalan, and 0.9% NaCl controls. Mice were treated with intraperitoneal injections and rats by ILP. TNF dosage was 20 microgram for mice and 200 microgram for rats. Melphalan was given at 5-10 mg/kg for both mice and rats. Results showed synergism of TNF and melphalan in both modes of therapy. In the systemic administration groups (mice carrying B16F10 and CT26 tumors), tumors increased in size in all but the combined TNF-melphalan group. In the regional delivery groups (rats carrying GF sarcoma cells treated via ILP), there was a 16% decrease in tumor volume in rats treated with TNF alone, a 29% decrease in rats treated with melphalan, and a 75% decrease in the combined TNF-melphalan group. In conclusion, TNF and melphalan proved to be highly synergistic in both systemic and regional delivery. This fact makes melphalan an adequate choice for TNF perfusion in advanced limb malignancies.

Animals↗
PubMed1996

Enforced c-KIT expression renders highly metastatic human melanoma cells susceptible to stem cell factor-induced apoptosis and inhibits their tumorigenic and metastatic potential.

Expression of the tyrosine-kinase receptor encoded by the c-KIT proto-oncogene progressively decreases during local tumor growth and invasion of human melanomas. To provide direct evidence that c-KIT plays a role in metastasis of human melanoma, we transfected the c-KIT gene into the c-KIT negative highly metastatic human melanoma cell line A375SM and subsequently analysed its tumorigenic and metastatic potential. A375SM parental cells, A375SM-NOT (neo, control), and A375SM-KIT-positive cells were injected s.c. and i.v. into nude mice. A375SM-KIT cells produced significantly slower growing s.c. tumors and fewer lung metastases than control cells. Exposure of c-KIT-positive melanoma cells in vitro and in vivo to stem cell factor (SCF), the ligand for c-KIT, triggered apoptosis of these cells but not of c-KIT-negative melanoma cells or normal melanocytes. Since SCF is produced by keratinocytes and other dermal cells in the skin, these results suggest that the loss of c-KIT receptor expression may allow malignant melanoma cells to escape SCF/c-KIT-mediated apoptosis, hence contributing to tumor growth and eventually metastasis. The antitumor and antimetastatic properties of SCF may be useful in treating human melanomas in early stages.

Animals↗
PubMed1996

Utilization of monensin for detection of microdomains in cholesterol containing membrane.

The effect of cholesterol on the monensin mediated proton-cation exchange reaction was measured in the time-resolved domain. The experimental system consisted of a black lipid membrane equilibrated with monensin (Nachliel, E., Finkelstein, Y. and Gutman, M. (1996) Biochim. Biophys. Acta 1285, 131-145). The membrane separated two compartments containing electrolyte solutions and pyranine (8-hydroxypyrene 1,3,6-trisulfonate) was added on to one side of the membrane. A short laser pulse was used to cause a brief transient acidification of the pyranine-containing solution and the resulting electric signal, derived from proton-cation exchange, was measured in the microsecond time domain. Incorporation of cholesterol had a clear effect on the electric transients as measured with Na+ or K+ as transportable cations. The measured transients were subjected to rigorous analysis based on numeric integration of coupled, non-linear, differential rate equations which correspond with the perturbed multi-equilibria state between all reactants present in the system. The various kinetic parameters of the reaction and their dependence on the cholesterol content had been determined. On the basis of these observations we can draw the following conclusions: (1) Cholesterol perturbed the homogeneity of the membrane and microdomains were formed, having a composition that differed from the average value. The ionophore was found in domains which were practically depleted of phosphatidylserine. (2) The diffusivity of the protonated monensin (MoH) was not affected by the presence of cholesterol, indicating that the viscosity of the central layer of the membrane was unaltered. (3) The diffusivity of the monensin metal complexes (MoNa and MoK) was significantly increased upon addition of cholesterol. As the viscosity along the cross membranal diffusion route is unchanged, the enhanced motion of the MoNa and MoK is attributed to variations of the electrostatic potential within the domains.

Arylsulfonates↗
PubMed1996

The mechanism of monensin-mediated cation exchange based on real time measurements.

Monensin is an ionophore that supports an electroneutral ion exchange across the lipid bilayer. Because of this, under steady-state conditions, no electric signals accompany its reactions. Using the Laser Induced Proton Pulse as a synchronizing event we selectively acidify one face of a black lipid membrane impregnated by monensin. The short perturbation temporarily upsets the acid-base equilibrium on one face of the membrane, causing a transient cycle of ion exchange. Under such conditions the molecular events could be discerned as a transient electric polarization of the membrane lasting approx. 200 microseconds. The proton-driven chemical reactions that lead to the electric signals had been reconstructed by numeric integration of differential rate equations which constitute a maximalistic description of the multi equilibria nature of the system (Gutman, M. and Nachliel, E. (1989) Electrochim. Acta 34, 1801-1806). The analysis of the reactions reveals that the ionic selectivity of the monensin (H+ > Na+ > K+) is due to more than one term. Besides the well established different affinity for the various cations, the selectivity is also derived from a large difference in the rates of cross membranal diffusivities (MoH > MoNa > MoK), which have never been detected before. (v) Quantitative analysis of the membrane's crossing rates of the three neutral complexes reveals a major role of the membranal dipolar field in regulating ion transport. The diffusion of MoH, which has no dipole moment, is hindered only by the viscose drag. On the other hand, the dipolar complexes (MoNa and MoK) are delayed by dipole-dipole interaction with the membrane. (vi) Comparison of the calculated dipoles with those estimated for the crystalline conformation of the [MoNa(H2O)2] and [MoK(H2O)2] complexes reveals that the MoNa may exist in the membrane at its crystal configuration, while the MoK definitely attains a structure having a dipole moment larger than in the crystal.

Arylsulfonates↗
PubMed1996

Protonation dynamics of the extracellular and cytoplasmic surface of bacteriorhodopsin in the purple membrane.

The dynamics of proton binding to the extracellular and the cytoplasmic surfaces of the purple membrane were measured by laser-induced proton pulses. Purple membranes, selectively labeled by fluorescein at Lys-129 of bacteriorhodopsin, were pulsed by protons released in the aqueous bulk from excited pyranine (8-hydroxy-1,3,6-pyrenetrisulfonate) and the reaction of protons with the indicators was measured. Kinetic analysis of the data imply that the two faces of the membrane differ in their buffer capacities and in their rates of interaction with bulk protons. The extracellular surface of the purple membrane contains one anionic proton binding site per protein molecule with pK = 5.1. This site is within a Coulomb cage radius (approximately 15 A) from Lys-129. The cytoplasmic surface of the purple membrane bears 4-5 protonable moieties (pK = 5.1) that, due to close proximity, function as a common proton binding site. The reaction of the proton with this cluster is at a very fast rate (3.10(10) M-1.s-1). The proximity between the elements is sufficiently high that even in 100 mM NaCl they still function as a cluster. Extraction of the chromophore retinal from the protein has a marked effect on the carboxylates of the cytoplasmic surface, and two to three of them assume positions that almost bar their reaction with bulk protons. The protonation dynamics determined at the surface of the purple membrane is of relevance both for the vectorial proton transport mechanism of bacteriorhodopsin and for energy coupling, not only in halobacteria, but also in complex chemiosmotic systems such as mitochondrial and thylakoid membranes.

Bacteriorhodopsins↗
PubMed1996

Quantitative evaluation of the dynamics of proton transfer from photoactivated bacteriorhodopsin to the bulk.

It has been reported by many research groups that protons released during the photocycle of bacteriorhodopsin are detected by surface bound indicators much faster than by indicators in the bulk. In this study we used numerical simulation of chemical reaction's dynamics for analyzing the delayed appearance of protons in the bulk. The results indicate that the low pK surface groups of the membrane, which form an undilutable concentrated matrix of proton binding sites, retain the protons in this space according to the mass action law. The main sites for proton accumulation are the cluster of carboxylates on the cytoplasmic side of the membrane. The protonation of an indicator in the bulk does not proceed by its reaction with free proton, but rather through self-diffusion of the indicator to the membrane and abstraction of proton from the protonated surface group. The detailed mechanisms which correspond with these reactions are reported.

Bacteriorhodopsins↗
PubMed1996

Regional perfusion with hemofiltration (chemofiltration) for the treatment of patients with regionally advanced cancer.

BACKGROUND: Regionally advanced cancer is a common, often unresolved problem. Effective local control with chemotherapy is limited by the toxicity following systemic administration. Chemofiltration (CF) is a form of regional perfusion that enables the administration of cytotoxic drugs into one body area while limiting systemic toxicity. The drug is infused into the artery supplying the involved area. The venous effluent of the same organ is pumped out into a hemofiltration unit at a high flow rate. The drug is then filtered away and the blood returned to systemic circulation. METHODS: Forty-one patients underwent 45 CF. Twenty-four patients had CF of the pelvis for advanced rectal carcinoma (10), malignant melanoma (6), and cancers of the uterine cervix (3), ovary (2), vulva (1), endometrium (1), and anus (1). Seventeen patients underwent CF of the liver for metastatic colon (10), breast (4), pancreas (1), ovary (1), and unknown primary (1) cancer. 5-fluorouracil (1 g/m2) and mitomycin-C (30 mg/m2); cisplatinum (200 mg/m2) alone or combined with bleomycin (50 mg/m2) and mitomycin-C (20 mg/m2); or melphalan (1 mg/kg) were the combinations used. RESULTS: Generally the procedure was well tolerated. Complications included transient leukopenia (18), paralytic ileus (2), hair loss (2), renal failure (1). Two patients died within 40 days following CF. Of 36 evaluable patients, 16 (44%) had partial response, 14 (38%) had stable disease, and 6 (18%) had disease progression. A decrease of at least 30% in carcinoembryonic antigen levels occurred in 12 of 24 patients (50%). Median time to progression was 7 months. Ten of 13 patients (77%) achieved good symptomatic palliation. CONCLUSIONS: The results of CF in our study are not superior to alternative methods of drug delivery to the liver and pelvis. However, considering that previous systemic chemotherapy had failed two-thirds of the patients, some benefit may be attributed to this regional delivery modality. Furthermore, pelvic CF afforded very significant symptomatic relief which was definitely superior to other methods.

Adult↗
PubMed1996

[Amplification of protooncogenes and expression of the HER2/neu oncogene in invasive breast cancer].

The protooncogene HER2/neu was analyzed in 27 women with invasive breast cancer and in 26 with benign breast conditions. Gene amplification was found in 3 patients, 1 in stage 3 and 2 in stage 4. In 3 patients in stage 2 overexpression of protein was found. Although amplification of the gene was found to be a significant predictor of clinical outcome (p < 0.05), high expression of the gene did not correlate with prognosis. In women with benign breast conditions there were no alterations of HER2/neu amplification may become a molecular marker in some malignancies. It should be noted that since few tumors show amplification, examination of a small series may not reveal clinical association with these alterations. Therefore, our conclusions require confirmation on a larger scale with long-term clinical follow-up.

Biomarkers, Tumor↗
PubMed1996

Increased microvascular permeability induced by prolonged interleukin-2 administration is attenuated by the oxygen-free-radical scavenger dimethylthiourea.

Effective use of interleukin (IL)-2 as an antineoplastic agent may be hindered by severe side-effects, in particular vascular leak syndrome, which leads to generalized, especially pulmonary, edema. The oxygen-free-radical scavenger dimethylthiourea (DMTU) was shown to attenuate IL-2-induced vascular leak syndrome in sheep receiving a single IL-2 injection. However, in the clinical setting multiple injections are necessary to gain a therapeutic effect. The present study tests whether DMTU attenuates IL-2-induced vascular leak syndrome following multiple IL-2 injections without affecting IL-2-induced cytotoxicity in peritoneal mononuclear cells. Mice were treated intraperitoneally with 1 x 10(5) units IL-2 three times daily for four consecutive days. DMTU (10 mg/0.5 ml) was administered to the study group once daily, prior to the first IL-2 injection. Comparing the wet/dry weight ratio of lungs, liver, and spleen showed that IL-2 caused a significant (P < 0.05) wet/dry increase in all three organs. DMTU attenuated the wet/dry increase in the lungs (P < 0.05), in the spleen (P < 0.05), and not at all in the liver. IL-2 induced a marked increase in peritoneal mononuclear cell counts, which was not attenuated by DMTU. The cytotoxic effect of IL-2-activated peritoneal mononuclear cells on target B16 cells was also unchanged in animals pretreated with DMTU. In conclusion, we have shown that DMTU ameliorates pulmonary permeability and vascular leak syndrome associated with multiple-dose IL-2 therapy, without eliciting an inhibitory effect on IL-2 induced-cytotoxicity.

Animals↗
PubMed1996

Interferon-beta prevents the upregulation of interleukin-8 expression in human melanoma cells.

The constitutive expression of interleukin-8 (IL-8) by human melanoma cells correlates with their metastatic potential. The exposure of human melanoma cells to the inflammatory cytokines IL-1 beta or tumor necrosis factor-alpha (TNF-alpha) upregulated IL-8 expression in a time-dependent and concentration-dependent manner. This enhanced expression of IL-8 was inhibited by cycloheximide or actinomycin-D. Treatment of melanoma cells with interferon (IFN) alpha, beta, or gamma did not affect the constitutive expression of IL-8, but IFN-alpha and IFN-beta blocked the upregulation of IL-8 expression in cells treated with IL-1 beta or TNF-alpha subsequent to or simultaneously with the IFN. These data suggest that the expression of IL-8 in human melanoma cells can be upregulated by inflammatory cytokines and that IFN-alpha and IFN-beta can counterregulate this stimulation.

Animals↗
PubMed1996

Intra-arterial chemotherapy with concomitant hemofiltration (chemofiltration) for treatment of loco-regional or pelvic metastases from malignant melanoma.

Loco-regional or pelvic metastases from malignant melanoma (MM) of the lower limbs or pelvis are usually refractory to systemic chemotherapy, the limiting factor being systemic toxicity. An attempt to improve this low response rate using a novel loco-regional approach involving intra-arterial high dose chemotherapy with concomitant hemofiltration of the venous effluent of the pelvis, hence chemofiltration, was studied. Chemofiltration was performed in seven MM patients. The arterial catheter and the venous cannula were placed in the aorta and the inferior vena cava just distal to the renal vessels. High-dose melphalan (1 mg/kg) or cis-platinum (200 mg/m2) was injected into the arterial catheter. Blood was pumped out into the hemofiltration unit at a rate of 500-700 ml/min. The filtered blood was returned via a catheter placed in the superior vena cava. Despite the extensive fluid exchange (9,700-15,000 ml), the procedure was well tolerated. Out of six patients who remained with measurable disease, three had a partial response lasting 5-12 months, two had stabilization of their disease for 3 months, and one developed a rapid progression. Chemofiltration is feasible in MM patients and is a viable option in locally advanced or metastatic malignant melanoma confined to the limb or pelvis.

Aged↗