Biomedical subjects

M Haba

Publications and source records attributed to M Haba.

12 recordsLinked to original sources

PubMed1998

[Pharmacokinetic analysis of scavenger receptor-mediated uptake of mucopolysaccharides in various cells].

Although the scavenger receptor-mediated uptake has been qualitatively investigated in the research fields of biochemistry and pathology, pharmacokinetic characteristics of the scavenger receptors are poorly understood. In this review, we summarized basic findings on scavenger receptors reported in available literature, and introduced our recent studies on the quantitative characteristics of the scavenger receptor-mediated uptake. High molecular weight fractionated [3H]heparin (HMWFH, 16,000-24,000 Da), one of the model mucopolysaccharides, was investigated to elucidate its uptake mechanism into isolated rat Kupffer cells, isolated peritoneal macrophages and liver parenchymal cells in primary culture. The equilibrium bindings of HMWFH to isolated Kupffer cells and peritoneal macrophages were concentration-dependent with the respective dissociation constants (Kd) of 5.7 and 6.0 nM and with the respective maximum binding capacities (Bmax) of 1.5 and 1.9 pmol/10(6) cells. Several ligands of scavenger receptors inhibited the binding of HMWFH to macrophages, suggesting the involvement of scavenger receptors in the uptake of HMWFH by these macrophages. It was also suggested that the scavenger receptor-mediated uptake is different from the receptor-mediated endocytosis of polypeptides and phagocytosis, based on the evidence of the now inhibitory effects of an inhibitor of receptor-mediated endocytosis of polypeptides(phenylarsine oxide) and phagocytosis inhibitors (cytochalasine B and colchicine) on the internalization. The involvement of scavenger-like receptors was also suggested in the uptake of HMWFH by liver parenchymal cells in primary culture by demonstrating inhibitory effects of ligands for scavenger receptors. The internalization into liver parenchymal cells by scavenger-like receptors was not affected by an inhibitor of receptor-mediated endocytosis of polypeptides and phagocytosis inhibitors, similarly to the results in the macrophage scavenger receptors. The Kd of 53.5 nM and Bmax of 32.8 pmol/10(6) cells in parenchymal cells were both in the order of magnitude larger than those in isolated Kupffer cells, suggesting the binding of HMWFH to scavenger-like receptors in parenchymal cells with lower affinity and higher capacity. On the other hand, an apparent internalization rate constant (kint, app) of 0.0056 min-1 was comparable with that in Kupffer cells (0.0118 min-1). Thus, we demonstrated the involvement of scavenger receptors in the uptake of HMWFH by rat Kupffer cells, peritoneal macrophages and liver parenchymal cells, and succeeded in characterizing the uptake kinetically. These findings should provide useful information for not only establishing the rational clinical use of mucopolysaccharides but also developing new drugs such as antiatherosclerotic agents and peptides delivered to cells with scavenger receptors.

Animals↗

PubMed1997

Kinetic characterization of binding and internalization of fractionated [3H]heparin in rat liver parenchymal cells in primary culture.

The binding and internalization of fractionated [3H]heparin (FH) was kinetically analyzed in rat liver parenchymal cells to clarify its cellular uptake mechanism. The binding of FH to the cell surface was saturable with the dissociation constant (Kd) of 53.5 nM and a maximum binding capacity (Bmax) of 19.9 pmol/mg protein. The binding of FH to the cell surface was competitively inhibited not only by heparan sulfate, a polyanion analogous to heparin, but also by rose bengal, an organic anion, suggesting the binding is based on an electric interaction requiring an anionic charge for substrates and consistent with the earlier suggestion of the involvement of the scavenger-like receptor. According to kinetic model analysis, the rate constants of association (K(on)), dissociation (k(off)), and internalization (k(int).app) were estimated to be 0.0005 nM-1 min-1, 0.0112 min-1, and 0.0056 min-1, respectively. Although both Kd and Bmax were larger than those reported in Kupffer cells, suggesting lower affinity and higher capacity in liver parenchymal cells, the apparent internalization rate constant was similar to that in Kupffer cells. We thus provided additional evidence suggesting that a scavenger-like receptor exists in rat liver parenchymal cells, and then kinetically characterized the surface binding and internalization of fractionated heparin by this receptor.

Animals↗

PubMed1996

Macromolecule-macromolecule interaction in drug distribution. V. Effects of plasma proteins on uptake of fractionated [3H]heparin in isolated rat Kupffer cells.

Effects of plasma proteins such as alpha-globulin on the uptake of high molecular weight (HMWFH: 23000 Da) and low molecular weight fractionated [3H]heparin (LMWFH: 10000 Da) were examined in isolated rat Kupffer cells. alpha-Globulin (8 mg/ml) affected neither surface binding nor internalization of LMWFH by Kupffer cells, while it reduced both surface binding and internalization of HMWFH without affecting the fraction internalized, which was a ratio of internalized amount to the total association. The total associations of HMWFH were about four times larger than that predicted assuming only the unbound fraction is available for uptake, suggesting the participation of protein-mediated transport in the uptake of HMWFH in Kupffer cells. Based on the same assumption, the saturable initial uptake of HMWFH versus concentration profile in the presence of alpha-globulin (8 mg/ml) was also analyzed to further examine the suggested protein-mediated transport. The estimated dissociation constant of 487 nM was three times larger than that in in vitro binding experiments (168 nM) and the binding capacity of 0.155 was one third of the value in vitro (0.5), suggesting apparent reductions in both binding affinity and capacity. Thus, we demonstrated the involvement of protein-mediated transport in the uptake of fractionated heparin in Kupffer cells and kinetically characterized it as the apparent enhancement of dissociation.

Alpha-Globulins↗

PubMed1996

Macromolecule-macromolecule interaction in drug distribution. IV. Molecular weight dependency in the interaction of fractionated [3H]heparin with plasma proteins.

Molecular weight dependency in the interaction of fractionated [3H]heparin (FH) with plasma proteins was evaluated by determining the protein binding of low molecular weight fractionated [3H]heparin (LMWFH: 7000 Da) and high molecular weight fractionated [3H]heparin (HMWFH: 16000 Da) by ultrafiltration and the effects of plasma proteins on the uptake in rat hepatocytes in primary culture. The unbound fractions of LMWFH were 0.5 and 0.8 in the presence of alpha-globulin and albumin, respectively, and were about 10 times larger than those of HMWFH, 0.04 and 0.1, suggesting a reduction in binding with a decrease in molecular weight. However, while the uptake of LMWFH was reduced by these proteins by the extents similar to bound fractions of LMWFH, the uptake of HMWFH was reduced by extents far smaller than bound fractions and comparable with those for LMWFH. Thus, it seemed that, while only unbound LMWFH is available for uptake, HMWFH bound to proteins is to some extent available for uptake (protein-mediated transport). The protein-mediated transport of heparin seemed to reduce with a decrease in molecular weight. It was also shown that the extended uptake of LMWFH was smaller than that of HMWFH not only in the absence of proteins but also in the presence of alpha-globulin, the major binding protein. The lower uptake of LMWFH is consistent with in vivo suggestion of lower hepatic accumulation.

Animals↗

PubMed1996

Uptake mechanism of fractioned [(3)H]heparin in isolated rat kupffer cells: involvement of scavenger receptors.

The uptake of fractionated [(3)H]heparin was examined to elucidate the uptake mechanism in isolated rat Kupffer cells. The equilibrium binding of fractionated [(3)H]heparin to Kupffer cells was concentration-dependent with the dissociation constant of 5.7 nM and the maximum binding capacity of 1.5 pmol/10(6) cells. Several ligands of scavenger receptors inhibited the binding of fractionated [(3)H]heparin to Kupffer cells competitively and also the internalization of heparin, suggesting the involvement of scavenger receptors in the uptake of fractionated [(3)H]heparin. Fractionated [(3)H]heparin was also suggested to be internalized according to first order kinetics with the apparent internalization rate constant of 0.010 min (-1). Lowering temperature from 37 to 4 degrees C reduced the fraction internalized from 33% to 6% without affecting the total association, while the fraction internalized at 25 degrees C was comparable with that at 37 degrees C. Metabolic inhibitors (2,4-dinitrophenol and rotenone), an inhibitor of receptor-mediated and adsorptive endocytosis of polypeptides (phenylarsine oxide) and phagocytosis inhibitors (cytochalasine B and colchicine) did not inhibit the internalization of fractionated [3(H)]heparin. As known inhibitors of receptor-mediated and adsorptive endocytosis of polypeptides and phagocytosis did not affect the uptake of fractionated heparin, the scavenger receptor-mediated uptake is suggested to be ATP-independent and different from receptor-mediated and adsorptive endocytosis of polypeptides and phagocytosis, although for temperature dependency it showed the typical characteristics of receptor-mediated endocytosis.

2,4-Dinitrophenol↗

PubMed1996

Molecular weight dependency in the uptake of fractionated [3H]heparin in isolated rat Kupffer cells.

The uptake of low molecular weight fractionated [3H]heparin (LMWFH, 10000 Da) was compared with that of high molecular weight fractionated [3H]heparin (HMWFH, 23000 Da) in isolated rat Kupffer cells. Several heparin analogs, including HMWFH and ligands of scavenger receptors, inhibited both the surface binding and internalization of LMWFH, suggesting the involvement of scavenger receptors in the uptake of LMWFH in isolated rat Kupffer cells as well as HMWFH, in spite of a large difference in molecular weight. Metabolic inhibitors (2,4-dinitrophenol and rotenone), receptor-mediated and adsorptive endocytosis of polypeptides (phenylarsine oxide) and phagocytosis inhibitors (cytochalasine B and colchicine) did not inhibit the internalization of LMWFH. These results suggest that the scavenger receptor-mediated uptake of LMWFH is ATP-independent and different from receptor-mediated and adsorptive endocytosis of polypeptides and phagocytosis, in agreement with our previous results for HMWFH. The equilibrium binding of LMWFH to Kupffer cells was concentration-dependent with the dissociation constant (Kd) of 50 nM and maximum binding capacity (Bmax) of 2.3 pmol/10(6) cells. The dissociation constant of LMWFH was an order of magnitude larger than that of HMWFH (5.7 nM), suggesting a decrease in binding affinity to scavenger receptors with a decrease in the molecular weight of fractionated heparin. It was also shown that LMWFH is internalized by scavenger receptors according to first-order kinetics with an apparent internalization rate constant (Kint,app) of 0.0053 min-1, which is about half that for HMWFH (0.0118 min-1). Molecular weight thus appears to be one of dominant factors determining the uptake of fractionated heparin by scavenger receptors in Kupffer cells, and may partly explain the reported lower hepatic uptake of low molecular weight heparin than that of unfractionated heparin.

2,4-Dinitrophenol↗

PubMed1995

Uptake of fractionated 3H-heparin by isolated rat Kupffer cells.

PURPOSE AND METHODS: The uptake of fractionated 3H-heparin by isolated rat Kupffer cells was examined to determine the uptake mechanism. RESULTS: The association of fractionated 3H-heparin was concentration-dependent with a dissociation constant of 3.4 nM and a maximum association capacity of 1.3 pmol/10(6) cells, suggesting the involvement of a specialized mechanism. Although 2,4-dinitrophenol inhibited neither the association nor internalization of fractionated 3H-heparin, lowering the temperature from 37 degrees C to 4 degrees C reduced the internalization of fractionated 3H-heparin by 70% without affecting the association. CONCLUSIONS: It is suggested that the uptake mechanism may differ from receptor-mediated endocytosis of polypeptides and be mediated by scavenger receptors, because organic anions, and several ligands of scavenger receptors, as well as several heparin analogs, inhibit the binding of fractionated 3H-heparin to Kupffer cells, while phenylarsine oxide, which is known to inhibit the receptor-mediated or absorptive endocytosis of polypeptides, inhibits neither the association nor internalization of fractionated 3H-heparin.

2,4-Dinitrophenol↗

PubMed1995

Uptake of low molecular weight fractionated [3H]heparin by rat hepatocytes in the primary culture.

The uptake of low molecular weight fractionated [3H]heparin (LMWFH: 7000 Da) was examined, for comparison with that of high molecular weight fractionated [3H]heparin (HMWFH:20000 Da), in a primary culture of rat hepatocytes. The uptake of LMWFH increased almost linearly with time up to 60 min (extended uptake), although a faster uptake was observed in the initial 2 min (initial uptake). Both the initial and extended uptake were saturable, and the maximum uptake velocity (Vmax) and the Michaelis constant (Km) were estimated to be 10.7 pmol/min/mg protein and 398 nM, respectively, for the initial uptake and 0.34 pmol/min/mg protein and 116 nM, respectively, for the extended uptake. The Km for the extended uptake was 5 times larger than that of 21 nM for HMWFH, but the other parameters were comparable with those for HMWFH. Thus, an increase in Km, or a decrease in the apparent affinity, with a decrease in molecular weight in the extended uptake may be responsible for the reported lower hepatic uptake of low molecular weight heparin, compared with unfractionated heparin. It was also shown that both the initial and the extended uptake of LMWFH were inhibited by several analogs of heparin, including HMWFH, and anionic compounds such as 4,4'-diisothiocyanatostilbene-2,2'-disulfonic acid (DIDS), suggesting that LMWFH and HMWFH, in spite of a large difference in the molecular weight, share the same specialized uptake mechanism, in which an anionic moiety and/or heparin-like structure plays an important role.

4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid↗

PubMed1993

Dose-dependent uptake of radioactivity by liver parenchymal and non-parenchymal cells after intravenous administration of fractionated 3H-heparin to rats.

The dose-dependent uptake of fractionated 3H-heparin in the subpopulations of liver cells, parenchymal and non-parenchymal cells, was characterized in rats in vivo. Following the intravenous administration of fractionated 3H-heparin, the radioactivity in plasma was eliminated according to the first order kinetics at each dose. However, the elimination rate constant decreased with dose over the dose range of 0.3 to 100 U/kg, suggesting nonlinear elimination. In accordance with the delay in the plasma elimination, the uptake rate constant of radioactivity by parenchymal as well as non-parenchymal cells of liver, the major distribution organ, also decreased. Although heparin has long been considered to be taken up by a reticuloendothelial system (RES) such as non-parenchymal cells in the liver, the uptake of fractionated 3H-heparin by parenchymal cells was found to be comparable with that by non-parenchymal cells at the lowest dose of 0.3 U/kg, and even larger than that by non-parenchymal cells at the highest dose of 100 U/kg. The uptake clearances of fractionated 3H-heparin at the dose of 0.3 U/kg were 86.4 and 504 ml/10(8) cells/d, respectively, for parenchymal and non-parenchymal cells. These values were much larger than those reported for polyvinylpyrrolidone, which has been suggested to be taken up by fluid phase endocytosis. Thus, the present study revealed the significant contribution of parenchymal cells in the hepatic uptake of fractionated 3H-heparin. The dose-dependent uptake with high clearance values in both parenchymal and non-parenchymal cells provides an in vivo suggestion of the specialized transport of fractionated heparin in these two subpopulations of liver cells.

Animals↗

PubMed1993

Macromolecule-macromolecule interaction in drug distribution. III. Kinetic characterization of the uptake of fractionated [3H]heparin and the effect of plasma proteins in the perfused rat liver.

The concentration-dependent hepatic uptake of fractionated [3H]heparin, a macromolecular model drug, was kinetically characterized and the effect of plasma proteins, albumin and alpha-globulin, was evaluated in the perfused rat liver as part of an ongoing effort to elucidate the mechanism of interaction of macromolecular drugs with biological macromolecules and the role of this interaction in the drugs' distribution. In the absence of proteins, the uptake of fractionated [3H]heparin was saturable with the maximum uptake velocity (Vmax) of 7.6 pmol/min/g liver and the Michaelis constant (Km) of 32.2 nM, suggesting the involvement of a specialized transport. alpha-Globulin (8.0 mg/ml) reduced the uptake of fractionated [3H]heparin at lower heparin at lower heparin concentrations. However, albumin (40 mg/ml) did not affect the uptake of fractionated [3H]heparin, suggesting an insignificant interaction. Assuming that fractionated [3H]heparin bound to alpha-globulin cannot be uptaken and that the reduction in uptake was solely attributable to the saturable Scatchard-type binding of fractionated [3H]heparin to alpha-globulin, the dissociation constant (Kd) and the binding capacity (n) were estimated to be 2.1 nM and 0.002, respectively. In in vitro binding experiments by ultrafiltration, Kd and n were estimated as 168 nM and 0.5, respectively, for alpha-globulin and 1021 nM and 0.02, respectively, for albumin, suggesting lower affinity and higher capacity in vitro for each protein.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1993

Uptake of fluorescein isothiocyanate (FITC)-fractionated heparin by rat parenchymal hepatocytes in primary culture.

The distribution of fractionated heparin in a primary culture of rat parenchymal hepatocytes was investigated optically using the fluorescence labelled drug and confocal imaging system with an inverted fluorescence microscope. The cell-associated fluorescein isothiocyanate (FITC)-fractionated heparin was observed to increase in conjunction with incubation time and also to localize, suggesting an internalization to cell organella with the exception of the nuclei.

Animals↗

PubMedDate not supplied

[Congenital malaria].

Several physiopathological hypothesis may explain the frequent transmission of malaria from a pregnant woman to the foetus, and its obstetrical consequences. Because of immunological reasons, such a transmission is most often silent, but some severe forms of congenital malaria do exist and they justify the chemoprophylaxy for pregnant women, and the treatment of any presumptive attack of malaria during pregnancy.

Antimalarials↗