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Biomedical subjects

V R Muzykantov

Publications and source records attributed to V R Muzykantov.

72 records · Page 4Linked to original sources
PubMed1987

In vivo administration of antibodies against type I collagen in rat: the specific accumulation in spleen.

[125I]-labelled rabbit antibodies against rat type I collagen and non-immune IgG were injected into rat circulation. The kinetics of their clearance and the biodistribution in different organs were studied. Both preparations showed very similar clearance rate, the kinetics fitting bi-exponential approximation with characteristic parameters t1,1/2 = 201 +/- 20 min before 320 min and t2,1/2 = 1350 +/- 450 min at times over 320 min for antibodies and 258 +/- 45 min and 890 +/- 140 min for IgG. The specific affinity of the circulating antibodies did not decrease within 24 hours. The antibodies were specifically accumulated in spleen, where their accumulation was 5-fold higher than that of non-immune IgG. Accumulation of antibodies was maximal 3 hours after the injection. The localization ratio (i.e. the ratio of the amount of the antibodies per g of tissue to that per g of blood) reached a maximum 24 hours after the injection and remained stable for 120 hours. Immunofluorescent staining of spleen sections resulted in a bright fluorescence of dense collagenous structures in the trabeculae and in the wall of the central follicular arterium, bright spot fluorescence in the marginal zone of the follicle, and diffuse fluorescence in the red pulp. These findings suggest an unusually high accessibility of collagen type I in spleen to circulating blood plasma components.

Animals↗
PubMed1987

Directed targeting of immunoerythrocytes provides local protection of endothelial cells from damage by hydrogen peroxide.

Red blood cells bearing anti-mouse IgG antibody on their surface (immunoerythrocytes) may provide for local protection of endothelial cells from the action of hydrogen peroxide. Subconfluent cultures of human umbilical vein endothelial cells responded sharply to increasing concentrations of hydrogen peroxide. Permeabilization of cellular membrane occurred at doses of hydrogen peroxide of from 1 to 3 mM, and was assured by incorporation of trypan blue stain immediately after treatment. Latent damage of cells produced by much lower doses of hydrogen peroxide (0.2-0.4 mM) could be observed after 24-hour incubation of treated cells in the normal culture medium with no hydrogen peroxide. The apparently dead cells differed from intact cells in morphology, were poorly attached to the substrate, and were readily incorporated by trypan blue, thus permitting easy visualization. Immunoerythrocytes bound to the antigen-coated surface enzymatically decreased the concentration of hydrogen peroxide in their microenvironment at least fivefold with respect to the total hydrogen peroxide concentration. Erythrocytes deposited on a part of the endothelial monolayer locally protected it from the damage at hydrogen peroxide concentrations ranging from 0.4 to 1.2 mM. Localization of protected zones corresponded precisely to the geometry of the erythrocyte coating. Immunoerythrocytes targeted to the endothelial cells by means of mouse anti-endothelial antiserum did not impair their viability and protected the endothelium from being killed at 0.3-1.2 mM hydrogen peroxide. This approach might be useful for a cell selection in mixed cell populations. The problem of local protection of cells involved in the inflammation focus are discussed.

Animals↗
PubMed1986

Immunotargeting of erythrocyte-bound streptokinase provides local lysis of a fibrin clot.

The creation of an anticollagen antibody-erythrocyte-streptokinase complex has been described. Immobilization of both proteins on erythrocyte membrane has been performed using an avidin-biotin interaction. Modification of streptokinase with (6-biotinylamido)hexanoic acid N-hydroxysuccinimide ester at the concentration of 1.1 mM (20% modification of protein amino groups) provides effective (up to 90%) attachment of streptokinase to an avidin-carrying erythrocyte surface. The loss of streptokinase activity due to modification under these conditions is not significant. The maximal attachment of streptokinase was equal to about 50 ng per 10(6) erythrocytes, i.e., about 5 X 10(5) molecules of streptokinase per erythrocyte. The presence of streptokinase in the incubation mixture inhibited the attachment of antibodies by about 50%. Nevertheless, co-immobilization of anticollagen antibody (1.0 X 10(5) molecules per cell) and streptokinase (2.8 X 10(5) molecules per cell) on the erythrocyte surface provided firm and specific binding of such erythrocytes to a collagen-coated surface (1.6 X 10(6) bound cells per 1 cm2 on a collagen-coated surface against 0.006 X 10(6) bound cells on a bovine serum albumin-coated surface). Targeting of such erythrocytes led to local lysis of a fibrin clot in the target zone. The properties described offer in principle the possibility of the application of this or a similar system of fibrinolytic agent targeting for the preventive therapy of rethrombosis during surgical manipulations on vessels.

Antibodies↗
PubMed1986

Red blood cell targeting to smooth muscle cells.

Monoclonal antibody discriminating between endothelial and smooth muscle cells is suggested to be used as a vector for directed transport of drugs to injured (denuded) areas of blood vessel wall. An in vitro model system was used in the studies: vascular smooth muscle or endothelial cells grown on plastic surface were treated with specific mouse monoclonal antibody recognizing an antigen localized on the surface of smooth muscle rather than endothelial cells; then erythrocytes coated with secondary (rabbit antimouse) antibodies were added. The results were analyzed spectrophotometrically or with scanning electron microscopy. Under the experimental conditions, erythrocytes, possible 'containers' for carrying the drugs, were found to bind only to smooth muscle cells. The data show that antibody provides absolute discrimination between endothelial and smooth muscle cells and, thus, may be used as a vector for drug targeting.

Animals↗
PubMed1986

Carrier-directed targeting of liposomes and erythrocytes to denuded areas of vessel wall.

Immunomorphological staining of specimens prepared from human carotid arteries with anti-collagen type I antibodies reveals large amounts of type I collagen in the subendothelium of lipid fibrous plaques. Collagen type I-containing structures, once in direct contact with blood after plaque rupture, can serve as potential targets for selective delivery of liposomes and erythrocytes to these areas. To verify this rationale, [14C]cholesterol oleate-containing liposomes were conjugated with bovine or human anti-collagen type I antibodies or human plasma fibronectin. Biotin derivatives of human anti-collagen type I antibody were coupled to human erythrocytes. Modified liposomes and erythrocytes were perfused in situ through segments of bovine, rabbit, or human arteries partially denuded with a balloon catheter prior to perfusion. After perfusion, the control and denuded areas were excised and subjected to scanning electron microscopic analysis and measurements of associated radioactivity. It was found that conjugates of liposomes or erythrocytes with anti-collagen type I antibodies or fibronectin are selectively bound by endothelium-free zones of arterial segments. Carrier-directed targeting of drug-laden liposomes and erythrocytes to thrombosis-prone areas of arterial lumen is discussed.

Animals↗
PubMed1986

[Targeted delivery of erythrocytes to human aortic smooth muscle cells].

Monoclonal antibodies specific for surface antigens of target cells are supposed to be good vectors for drug transport. It is suggested using monoclonal antibodies that distinguish between smooth muscle and endothelial cells as vectors for directed drug transport to injured (denuded) areas of the blood vessel wall. The following in vitro model was used: monoclonal antibodies were added to cultured vascular smooth muscle or endothelial cells, this was followed by the addition of erythrocytes conjugated with rabbit antimouse antibodies. Spectrometry and scanning electron microscopy were used to assess the results. The erythrocytes, possible containers of drugs, under the experimental conditions were found to bind only to smooth muscle cells. The data obtained suggest that antibody IIG10 discriminating between smooth muscle and endothelial cells provides a specific tool for erythrocyte delivery to smooth muscle cells.

Animals↗
PubMed1985

Evaluation of quantitative parameters of the interaction of antibody-bearing liposomes with target antigens.

The model system for the analysis of targeted liposomes is proposed--the layer of protein antigen adsorbed on polystyrene wells. Antibodies were treated with palmitoyl chloride and liposomes were produced by the cholate dialysis method in the presence of the modified protein (7 X 10(-4) mol protein/mol lipid). Affinity of antibody-bearing liposomes to the antigen on the surface of Multiwell plates was studied, and apparent dissociation constant value was estimated: KD was in the range 1.5 to 5 X 10(-9) M liposomes. Sequential transfers of liposomes in antigen-coated plates revealed that the high-affinity fraction of liposomes is adsorbed first. The bound fraction has 1.7-times-higher protein content. For effective in vivo targeting it would be necessary to have high-affinity liposomes and a high concentration of the target antigen.

Animals↗
PubMed1985

Targeting of enzyme immobilized on erythrocyte membrane to collagen-coated surface.

It is suggested to use 'enzyme(s)-erythrocyte-antibody' complex for modulation of the microenvironment in definite compartments of blood circulation. A model system including peroxidase, human erythrocytes and anti-collagen antibodies was chosen to illustrate the principle. Peroxidase was conjugated to the erythrocyte surface via periodate-oxidized enzyme carbohydrate moiety; biotinylated antibodies were linked by avidin to the biotinylated erythrocytes. The properties of the immunocomplexes obtained have been investigated in an artificial system simulating an injured blood vessel wall. The advantages in using erythrocyte-mediated immunoenzyme complexes for enzyme (drug) targeting are discussed.

Antibodies↗
PubMed1985

Distribution of type I, III, IV and V collagen in normal and atherosclerotic human arterial wall: immunomorphological characteristics.

35 autopsies--aged 30 to 75 years--were investigated in order to establish trends of collagen localization in various types of arteries depending on age, arterial size and degree of atherosclerosis. Cryostat sections stained with highly specific antibodies to human types I, III, IV or V collagen, or with the antiserum to smooth muscle myosin were examined by the indirect immunofluorescence technique. Localization of type III collagen was very similar to that of type I. Fibrous structures of both type I and type III were then major constituents of the intima, media and adventitia. Sparse fibrils of type I and type III collagens were revealed in the subendothelium of unaffected intima. They gradually became abundant in the deeper intimal layers contrasting with loose fibrillar formations of the media. The content of interstitial collagens was significantly increased in the subendothelium of local intimal thickenings and in a thickened intima of the aged. This fact, considering the thrombogenicity of interstitial collagens, may be relevant to the atherogenesis through the "response-to-injury" mechanism. Type IV and type V collagens are localized to the endothelial basement membrane and basement membranes of smooth muscle cells of the intima and media. Diffusely distributed type V collagen was also observed in the intercellular space of the intima. In lipid streaks, parallel layers of condensed interstitial collagens separated groups of cells and extracellular lipid depositions. In fibrous plaques, types I and III became prevalent structural elements and their densely packed fibers occupied whole regions devoid of any type IV and type V collagen. Heavily thickened type IV collagen structures surrounding individual smooth muscle cells were found in fibrous plaques, but never, in unaffected intima.

Adult↗
PubMed1985

[Localization of different types of collagen (I, III, IV, V) in the connective tissue of the popliteal artery and skeletal muscle of man (immunoelectronmicroscopic analysis)].

By means of immunoperoxidase and immunoferritin techniques collagen of the I, III, IV and V types has been revealed in cryostat sections of the popliteal artery and in the musculus quadriceps of the femur. Areas of the vascular wall without any macroscopical signs of lesions have been investigated. They have been obtained from amputated extremities of young persons (17-22 years old), and muscle pieces have been taken during operations performed in the knee joint. After certain immunocytochemical procedures the cryostat slices are embedded in mixture of epon 812 and araldit, non-contrasted ultrathin slices are examined in the electron microscope JEM 100CX. Collagen of the I and III types is revealed in fibrills 20-80 nm thick either with or without cross striation, as well as in microfibrills. Collagen of the III type in the intercellular substance of the arterial wall occurs in nonfibrillar form. Collagen of the IV type is revealed in basal membranes of the smooth muscle cells of the arterial wall, of the muscle fibers and of endothelium of blood capillaries of the skeletal muscle. Collagen of the V type is found as accumulations having various size and form; they localize in many places of the intercellular substance of the arterial wall. A tight contact is revealed between the formations including collagen of the V type with drops of elastin and elastic fibers. A suggestion is made that collagen of the V type participates in formation of elastic fibers.

Adolescent↗
PubMed1985

Hemolytic complement activity assay in microtitration plates.

A new rapid technique is developed for the determination of complement activity in a large number of samples. Following serial dilution of complement, hemolysis is performed in the same microtiter plate. After the reaction, the degree of hemolysis in wells of the plate is determined spectrophotometrically by measurement of "absorbance" (light scattering) at 630 nm, without additional procedures. This method can find application in clinical and experimental biochemistry for the analysis of a large (up to thousands) number of samples.

Complement System Proteins↗
PubMed1983

Type I and III collagens as a possible target for drug delivery to the injured sites of vascular bed.

Interaction of anti-human collagen types I and III antibodies, as well as human red blood cells conjugated with these antibodies, with the surface of denuded intima of human aorta has been studied. Data on the accessibility of antigenic determinants of collagen types I and III for antibodies and red blood cells conjugated with these antibodies have been obtained in ex vivo experiments in an original model. On the basis of the obtained results it is concluded that antigenic determinants of collagen types I and III exposed as a result of blood vessel wall injury can serve as a target for drug delivery to the injured site(s).

Antibody Specificity↗
PubMed1983

Red blood cell targeting to collagen-coated surfaces.

The interaction of human red blood cells carrying antihuman collagen antibody with collagen-coated surfaces was studied. Avidin was used as bifunctional crosslinking agent for the attachment of antibody to the red blood cell surface. Antibody-carrying red blood cells efficiently and specifically bound to collagen-coated surface covering a significant part of the surface. The components of normal blood had an insignificant effect on red blood cell binding. A model of drug targeting to the injured sites(s) of blood vessel wall is proposed.

Antibody Specificity↗
PubMedDate not supplied

Modulation of angiotensin-converting enzyme in cultured human vascular endothelial cells.

Previous work has suggested that not all immunoreactive angiotensin-converting enzyme (ACE) in tissues or cells is in a biologically active state. We have explored this possibility in cultured human umbilical vein endothelial cells (HUVEC), one of the most widely studied in vitro endothelial cell systems. Our approach included characterization of the effect of increasing passage number on ACE activity and expression of immunoreactive ACE at the single cell level, the subcellular compartmentalization of active ACE, and the effect of phorbol ester (PMA) treatment. We found that both ACE activity and expression of ACE antigen were downregulated by cultivation (30% of ACE-positive cells at seventh passage vs. 90% in primary culture). ACE downregulation is specific (number of CD31-positive cells did not change with cultivation) and correlated with downregulation of factor VIII-antigen. The percentage of ACE-positive cells in permeabilized HUVEC at third passage was almost twice that in nonpermeabilized HUVEC (90% vs. 50%), indicating that HUVEC contain intracellular immunoreactive ACE. ACE activity, however, was similar when measured in intact cells and in cell lysates. Moreover, diazonium salt of sulfanilic acid (DASA), a membrane-impermeable ACE inhibitor, inhibited ACE activity in intact cells and in cell lysates at the same extent, thus implying that intracellular ACE is inactive. PMA (100 nM) treatment increased the percentage of ACE-positive cells at third passage from 57 to 96%. ACE activity was increased 3-fold in cell and 1.5-fold in the culture medium of PMA-treated cells. Analysis of ACE activity in intact monolayers and cell lysates of control and PMA-treated cells revealed that all enzymatically active ACE in PMA-treated cells is localized on the plasma membrane and acts as an ectoenzyme. We conclude that expression of ACE by HUVEC is downregulated by repeated passage in culture but can be restored by PMA treatment. In addition, ACE expression is heterogeneous between neighboring cells, and total immunoreactive ACE protein associated with HUVEC includes an inactive pool of the enzyme.

Antigens, Surface↗
PubMedDate not supplied

Effect of flow rate and blood cellular elements on the efficiency of red blood cell targeting to collagen-coated surfaces.

The effect of bloodstream factors (flow rate and blood cellular elements) on the behavior of an experimental system simulating drug targeting to injured sites of vessel walls was studied. The system consisted of red blood cells carrying antibody to type I human collagen (drug carrier) and plastic tube with a collagen-coated inner surface (target). The tube was perfused with a 0.5% (v/v) suspension of 51Cr-labeled red blood cells at different linear flow rates and the red blood cell binding to the tube was determined by gamma-counting. It was demonstrated that an increase in the linear flow rate from 0 to 2 cm/s leads at first to increase of red blood cell binding from 2 X 10(5) to 7 X 10(5) cells/cm2 and then to the decrease of binding back to 2 X 10(5) cells/cm2. In the presence of 50% (v/v) of intact red blood cells the binding continuously increases from 2 X 10(5) to 2.5 X 10(6) cells/cm2 without a subsequent drop. On the basis of the obtained results it is concluded that the behavior of the systems for drug targeting in simple in vitro models can drastically differ from the conditions present in vivo.

Antibodies↗