Characterization of a 95 kDa high affinity human high density lipoprotein-binding protein.

A new human 95 kDa high density lipoprotein (HDL)-binding protein (HBP) corresponding to a high affinity HDL-binding site with K(d) = 1.67 microg/mL and a capacity of 13.4 ng/mg was identified in human fetal hepatocytes. The HDL binding with the 95 kDa HBP plateaus at 2.5-5 microg/mL under reducing and nonreducing conditions. The association of HDL(3) with the 95 kDa HBP plateaued in 15-30 min while dissociation was complete in 30 min. HDL(3), apoA-I, and apoA-II were recognized by the 95 kDa HBP while low density lipoproteins (LDL) and tetranitromethane-modified HDL were not. The 95 kDa HBP predominantly resides on the surface of cells since trypsin treatment of HepG2 cells eliminated nearly 70% of HDL binding. All studied human cells and cell lines (HepG2, Caco-2, HeLa, fibroblasts, SKOV-3, PA-I) demonstrated the presence of the 95 kDa protein. Both RT-PCR and Western blotting for HB-2/ALCAM were negative in human fetal hepatocytes while Gp96/GRP94 was clearly differentiated from the 95 kDa HBP by two-dimensional electrophoretic mobility. Moreover, deglycosylation of HepG2 membrane preparations did not affect either HDL binding to the 95 kDa HBP or its size, while in contrast it affected the molecular weights of HB-2/ALCAM and SR-BI/CLA-1. We conclude that the 95 kDa HBP is a new HDL receptor candidate widely expressed in human cells and cell lines.

Heat shock protein 60 is a high-affinity high-density lipoprotein binding protein.

A new 55-kDa HDL/apolipoprotein binding protein was demonstrated in plasma membrane preparations of the human cell lines and primary cultured hepatocytes. Analysis of specific binding by ligand immunoblots of HDL, apoA-I, and apoA-II to a partially purified 55-kDa PA-I plasma membrane preparation demonstrated a K(d,HDL) = 50 nM (10 microg/ml), K(d,apoA-II) = 20 nM (0.4 microg/ml), and K(d, apoA-I) = 330 nM (10 microg/ml). Following preparative SDS-PAGE electrophoresis of a plasma membrane preparation isolated from human PA-I cells, fractions with apoA-II binding activity were collected, concentrated, and subjected to two-dimensional electrophoresis. Internal microprotein sequencing of the 55-kDa protein band revealed the binding protein as being heat shock protein 60 (hsp60). The hsp60 monoclonal antibody LK-1 blocked apoA-II binding to the 55-kDa HBP preparation. In summary, these results provide a potential mechanism to explain the known association between immunity developed against hsp60 and the development of atherosclerosis.

[Characteristics of the strategy of intensive care of unconscious children]. / Osobennosti reanimatsionnoi taktiki u detei v bessoznatel'nom sostoianii.

Three major groups can be distinguished among children hospitalized in resuscitation and intensive care wards in a state of unconsciousness: patients with bacterial meningitis and meningoencephalitis, viral meningoencephalitis, and noninfectious involvement of the NCS. Common strategy of treatment of these patients is characterized by some specific features: etiotropic antibiotic therapy in group 1 patients, antiviral drugs in group 2, and mainly symptomatic measures in group 3. Immunotherapy used in each of these groups is substitute in group 1, immunomodulating and immunostimulating in group 2 and even more so in group 3. Better results in group 1 are due to highly effective etiotropic antibiotics and drugs.

[Diagnostic value of several immunity parameters in gestoses]. / Diagnosticheskaia tsennost' nekotorykh pokazatelei immuniteta v klinike gestozov.

Gestosis is a grave complication of pregnancy. Assessment of its severity in various pregnancy terms is important for making decision about preterm delivery and predicting the outcome of pregnancy. The authors consider that such immunological tests as measurement of circulating immune complexes, NBT test, and cytological index of activity are reliable indicators of the severity of gestosis, which can be used for predicting the condition and its development.

[The effect of insulin on protein synthesis and secretion in primary liver cultures]. / Vliianie insulina na sintez i sekretsiiu belkov v pervichnykh kul'turakh kletok pecheni.

The effects of insulin on the rates of synthesis of intracellular and secreted proteins have been studied in primary cultures of fetal rat hepatocytes. During 3-h incubation with the hormone and 14C-L-leucine similar stimulation of synthesis of both intracellular and secreted protein was observed. Insulin did not affect significantly the proportion of 14C-labelled proteins destined for secretion in the cell cultures grown on standard nutrient medium, as well as on selective medium containing phenobarbital and cortisol. The results obtained show that the major part (85-90%) of newly synthesized proteins remains inside the cells. Therefore, biological testing of insulin or its synthetic analogs on hepatocyte cultures may be limited by the estimation of the rate of biosynthesis of intracellular proteins only.

[The possible participation of gap junctions in the realization of the effects of stimulators of secretory processes in the hypophysis]. / Vozmozhnoe uchastie shchelevykh kontaktov v realizatsii éffektov stimuliatorov sekretornykh protsessov v gipofize.

Sodium butyrate and octanol did not change the basal rate of GH secretion. However, octanol completely and partially suppressed GH release stimulated by thyroliberin and DbcAMP, respectively. Octanol did not influence GH secretion induced by DbcGMP. Besides, octanol did not change significantly basal prolactin release, but this agent blocked secretogenic action of on lactotrophs. The results show the important role of cell-to-cell communication mediated by gap junctions in stimulatory action of thyroliberin and cAMP analogue on GH release from neonatal pituitary cells, as well as in secretogenic action of thyroliberin on lactotrophs.

[Reactions of rat fetal hepatocytes to hormones in primary cultures grown in selective media with an addition of glucocorticoid and barbiturate]. / Reaktsii gepatotsitov plodov krys na gormony v pervichnykh kul'turakh vyrashchennykh v selektivnoi srede s dobavleniem gliukokortikoida i barbiturata.

After growing of fetal rat hepatocytes in the medium containing cortisol and sodium barbital for 4-8 days and subsequent cultivation in the medium without glucocorticoid and barbiturate for 1-4 days insulin retained its ability to stimulate total RNA and protein synthesis, while human growth hormone kept its enhancing action on RNA biosynthesis. However, cortisol did not change protein synthesis and inhibited paradoxically incorporation of 3H-uridine into total RNA, after preincubation of cells in the selective medium. This suggests that prolonged exposure of cultured fetal rat liver cells to cortisol and sodium barbital may cause phenomena similar to those of hormonal and/or enzymatic imprinting.

[Effects of dibutyryl derivatives of cyclic nucleotides on synthesis of total RNA and proteins in cultured fetal rat hepatocytes]. / Vliianie dibutirilproizvodnykh tsiklicheskikh nukleotidov na sintez summarnykh RNK i belka v kul'tiviruemykh gepatotsitakh plodov krys.

During short-term (6 h) or long-term (24 h) incubation of fetal rat liver cells in primary cultures, 10(-3) M dibutyryl-derivative of cyclic AMP (Bt2cAMP) and sodium butyrate decreased total RNA and protein synthesis. In contrast, dibutyryl-cyclic GMP (Bt2cGMP) at the same dose (10(-3) M) was without significant effect on RNA and protein biosynthesis. During short-term (4 h) incubation 10(-3) M Bt2cAMP and Bt2cGMP stimulated serum albumin production, while sodium butyrate was without effect. In long-term (22 h) incubation only 10(-3) M Bt2cAMP noticeably increased albumin production. The results obtained clearly show that Bt2cGMP, unlike Bt2cAMP, is not able to modify significantly total RNA and protein synthesis in cultured fetal rat liver cells. It is concluded also that the effects of dibutyryl-derivatives of cyclic nucleotides, at least on albumin production, are not mimiebet by butyrate.

[The direct stimulating action of thyroliberin on the biosynthesis of total protein in cultured liver cells of rat fetuses]. / Priamoe stimuliruiushchee vliianie tiroliberina na biosintez summarnogo belka v kul'tiviruemykh kletkakh pecheni plodov krys.

In experiments using fetal rat liver cultured cells TRH was shown to stimulate total protein synthesis but not RNA synthesis during long incubation. Somatostatin affected neither protein synthesis nor RNA synthesis in cultured liver cells. Possible physiological role of peripheral TRH during perinatal period in the rat is discussed.

[Hormonal regulation of the production of serum albumin by cultured hepatocytes of rats during prenatal and postnatal period of development]. / Gormonal'naia reguliatsiia produktsii syvorotochnogo al'bumina kul'tiviruemymi gepatotsitami krys v pre- i postnatal'nom periode razvitiia.

The effects of several hormones on the production of immunoreactive serum albumin (SA) were examined in primary cultures of liver cells obtained from rat fetuses on 21-22 days of gestation of from 3-week old rats. Cortisol, bovine insulin and human growth hormone stimulated SA production in both types of liver cell cultures during 20 h-incubation. L-Triiodothyronine (T3; 10(-9)-10(-7) M) weakly stimulated SA production by hepatocytes from the rats, but markedly inhibited it in cultures of fetal rat liver cells in a dose-dependent manner. In contrast, T3 action on total RNA and protein biosynthesis, estimated as the incorporation of labelled precursors into macromolecules, was stimulatory one in both types of cell cultures. It is concluded that hormonal regulation of SA production is similar in cultured liver cells from fetal and early postnatal rats except for the action of T3. The physiological importance of striking developmental change of T3 action on SA production remains to be determined.

[The effect of dibutyryl derivatives of cyclic nucleotides on albumin production in a primary culture of rat liver cells]. / Osobennosti vliianiia dibutirilproizvodnykh tsiklicheskikh nukleotidov na produktsiiu al'bumina v pervichnykh kul'turakh kletok pecheni krys.

Dibutyryl-derivative of cyclic AMP (Bt2cAMP, 10(-3)M) affected the basal and cortisol-induced production of immunoreactive serum albumin (SA) in primary cultures of fetal rat liver cells by the type of two-phase time dependency. The same two-phase character exhibited dose-dependent effect of Bt2cAMP (10(-5)-10(-3)M) on SA production in cultured liver cells obtained from fetal and 3 weeks old rats. Similar two-phase dose- and time-dependent effects demonstrated Bt2cGMP (10(-5)-10(-3)M), although certain differences were detected when either action of Bt2cAMP and Bt2cGMP or response of cultured liver cells from fetal and early postnatal rats to these drugs were studied. The data obtained suggest that normal liver cells are provided with feed-back mechanism which limited the cellular reaction to cyclic nucleotides under experimental conditions involving, potentially excessive doses and/or time of action of these agents.

[Hormonal regulation of total RNA and protein biosynthesis in liver cell cultures of rats in the pre- and postnatal period of development]. / Gormonal'naia reguliatsiia biosinteza summarnykh RNK i belka v kul'ture kletok pecheni krys pre- i postnatal'nogo perioda razvitiia.

The effects of several hormones on total RNA and protein biosynthesis were examined in primary cultures of liver cells obtained from rat fetuses on 21-22 days of gestation and from 3 week-old weanling rats. The intensity of biosynthesis processes was estimated by the incorporation of labeled precursors in macromolecules. Insulin, cortisol, triiodothyronine (T3) stimulated RNA and protein biosynthesis in both types of cultures. These hormones enhanced total protein biosynthesis in fetal rat liver cells more efficiently than in hepatocytes of weanling rats. Somatotropin (growth hormone--GH) did not change total protein biosynthesis but notably increased RNA synthesis and the production of immunoreactive serum albumin. Experiments on fetal rat liver cell cultures showed that stimulating action of cortisol on RNA synthesis was synergistic in relation to the effects of insulin and GH. It has been concluded that fetal rat liver cell at the end gestation are able to respond adequately to anabolic action of the hormones.