The relationship of the antepartum elevation in serum relaxin levels in pregnant rats to luteolysis was examined by determining the effects of the luteolysin prostaglandin F2α (PGF2α) and the prostaglandin synthetase inhibitor indomethacin on antepartum serum relaxin levels, as well as on luteolysis and birth. Intravenous administration of PGF2α on the morning of Day 20 elevated serum relaxin levels approximately fourfold within 15 min. Administration of the prostaglandin synthetase inhibitor indomethacin from Day 19 until Day 23 protracted luteolysis, delayed or prevented birth, and delayed the antepartum elevation of serum relaxin levels, until after indomethacin treatment had been terminated. Collectively, these results indicate that prostaglandins, in particular PGF2α, may promote the antepartum increase in serum relaxin levels, as well as luteolysis and birth in rats.
Two monoclonal antibodies designated BAT085 and G3-136 were raised by immunizing BALB/c mice with gp120 purified from human immunodeficiency virus type 1 (HIV-1) IIIB-infected H9 cell extracts. Among three HIV-1 laboratory isolates (IIIB, MN, and RF), BAT085 neutralized only IIIB infection of CEM-SS cells, whereas G3-136 neutralized both IIIB and RF. These antibodies also neutralized a few primary HIV-1 isolates in the infection of activated human peripheral blood mononuclear cells. In indirect immunofluorescence assays, BAT085 bound to H9 cells infected with IIIB or MN, while G3-136 bound to H9 cells infected with IIIB or RF, but not MN. Using sequence-overlapping synthetic peptides of HIV-1 IIIB gp120, the binding site of BAT085 and G3-136 was mapped to a peptidic segment in the V2 region (amino acid residues 169 to 183). The binding of these antibodies to immobilized gp120 was not inhibited by the antibodies directed to the principal neutralization determinant in the V3 region or to the CD4-binding domain of gp120. In a competition enzyme-linked immunosorbent assay, soluble CD4 inhibited G3-136 but not BAT085 from binding to gp120. Deglycosylation of gp120 by endo-beta-N-acetylglucosaminidase H or reduction of gp120 by dithiothreitol diminished its reactivity with G3-136 but not with BAT085. These results indicate that the V2 region of gp120 contains multiple neutralization determinants recognized by antibodies in both a conformation-dependent and -independent manner.
A novel treatment has been devised in our studies of the purification of inhibin from porcine and human follicular fluids (pFFl and hFFl, respectively). Both pFFl and hFFl were precipitated with acetone and extracted with acetic acid to provide a starting material for subsequent gel filtration and reverse-phase high-pressure liquid chromatography (HPLC). Inhibin from pFFl was purified 4200-fold using this methodology. Inhibin from hFFl could not be purified to this degree since recoveries were relatively poorer than for pFFl and yielded too little material for the HPLC step.
Relaxin and progesterone are produced by the corpora lutea in the pregnant rat. Relaxin immunoactivity levels are elevated in peripheral sera during the last 12 days of pregnancy. In rats maintained under a conventional photoperiod of 14 h of light (0600–2000 h) and 10 h of darkness (2000–0600 h), there is an antepartum elevation in serum relaxin to maximal levels coincident with the rapid decline in serum progesterone to basal levels during the light phase of the photoperiod 1 day before birth. Therefore, we postulated that this maximal elevation in serum relaxin levels may be temporally associated with functional luteolysis and linked to the photoperiod. In the present study the photoperiod was advanced near midpregnancy in order to examine further the relationship of the antepartum elevation in serum relaxin levels with both functional luteolysis and the photoperiod. Three groups of rats were maintained under a conventional photoperiod of 14 h of light (0500-1900 h) and 10 h of darkness until days 7 and 8 of pregnancy when the photoperiod was advanced 8 h in group 2 (G2) and advanced 18 h in G3 relative to the conventional photoperiod that was maintained in Gl. Serum relaxin and progesterone levels were determined in blood samples obtained at 4-h intervals from 2000 h on day 19 of pregnancy until birth. The times of occurrence of birth, maximal relaxin levels, and decline of progesterone to basal levels in G2 and G3 were generally advanced 50–60% of the advancement of the photoperiod. There was a close temporal association between the attainment of maximal relaxin levels and basal progesterone levels; they both occurred during the light phase of the photoperiod, approximately 24 h before birth in all three groups. We conclude that the antepartum elevation of serum relaxin to maximal levels may be associated with functional luteolysis and that its time of occurrence is influenced by the photoperiod. This study also provides evidence that the antepartum elevation of relaxin levels consists of two phases which occur at a 24-h interval. It is proposed that these two phases in the elevation of relaxin levels may be indicative of an increasingly effective endogenous circadian luteolytic process whose time of occurrence is influenced by the light-dark schedule. (Endocrinology113:997, 1983)
The role of the maternal pituitary gland and LH in the regulation of the release of relaxin from the onset of the light phase on day 21 of pregnancy (day 21) until delivery (herein designated the antepartum period) was examined in rats. Serum relaxin levels were determined in rats hypophysectomized (HYPOX) or sham hypophysectomized (SHAM) on day 14. In SHAM rats, the elevated serum relaxin levels that occurred during the light phase of days 21 and 22 coincided with a rapid decline in serum progesterone to basal levels, and relaxin levels then declined markedly throughout the 18- to 24-h before delivery. During the antepartum period HYPOX rats differed from SHAM rats in the following ways: 1) mean serum relaxin levels were higher immediately before the antepartum period (day 20) and remained at the maximal levels the sham-operated animals experienced throughout the ensuing antepartum period; 2) elevations in relaxin levels generally occurred in individual rats; however, they usually occurred later than day 22 and, in many cases, during the dark phase of the photoperiod; 3) mean relaxin levels remained elevated throughout days 22 and 23; and 4) the antepartum decline in progesterone levels was protracted throughout and beyond day 22. Subsequent experiments determined the effects of LH on birth, relaxin levels, and functional luteolysis. Some Sprague-Dawley-derived rats give birth during the light phase of day 22 (day 22 rats), but generally, most give birth during the light phase of day 23 when maintained under the housing conditions employed for these studies. When LH was injected to both intact and HYPOX rats in the delay-vehicle polyvinylpyrrolidone on the morning of day 20, birth and antepartum ovarian events were advanced. All intact rats and all but one HYPOX rat gave birth on day 22, and the time of occurrence of the rapid decline in both serum relaxin and progesterone levels coincided with these events in day 22 rats. In contrast, a reduction of endogenous serum LH levels in intact rats by means of the injection of a LH antiserum from days 19 through 22 delayed birth as well as the antepartum decline in both serum relaxin and progesterone levels. The results of this study indicate that maternal pituitary LH may play a role in promoting the normal progression of birth, the antepartum elevation and decline in serum relaxin levels, and functional luteolysis in the pregnant rat.
Therapeutic approaches that interfere with viral entry hold promise in preventing or treating HIV infection. Hu5A8, a humanized monoclonal antibody against CD4, was previously shown to inhibit HIV and SIV replication in vitro and was safely administered to rhesus monkeys without depleting CD4+ T cells. This antibody completely suppressed replication of six different SIVmac 251 primary isolates in vitro. Twice weekly administration of 3-mg/kg doses of hu5A8 for 2 to 4 weeks to SIV-infected rhesus monkeys resulted in sustained plasma antibody levels of ≥20 μg/ml during treatment and 5- to 50-fold decreases in plasma viremia, although suppression of viral replication was transient. Two of three treated monkeys developed antibody responses against the administered monoclonal antibody. Loss of antiviral effect was not temporally associated with anti-hu5A8 antibody responses or due to activation of CD4+ T cells by hu5A8. However, SIV isolated after hu5A8 treatment was approximately 5-fold more resistant to suppression by hu5A8 than SIV isolates obtained from the same monkeys before treatment. The rapid development of resistance may have resulted from SIV variants that infect cells by a CD4-independent mechanism. These results support the overall concept of anti-CD4 monoclonal antibody treatment to suppress AIDS virus replication in vivo while demonstrating important issues as to its clinical feasibility.
Murine mAb BAT123, which was made against the envelope glycoprotein gp120 of HTLV-IIIB strain of HIV type 1 (HIV-1), is capable of neutralizing HTLV-IIIB in vitro. It also inhibits the fusion between uninfected CD4+ cells and HIV-1-infected cells to form syncytia. As a step to explore the potential utility of the anti-HIV antibody in vivo, we have constructed a mouse-human chimeric antibody by rDNA techniques. The chimeric antibody, which bears the variable domains of mouse antibody BAT123 and constant domains Cr1 and C kappa of human Ig retains the Ag specificity of BAT123 as determined by its reactivity with HIV-1-infected H9 cells, gp120 in Western blot analysis, and the oligopeptide recognized by BAT123. The antiviral activities of the chimeric antibody in neutralizing HIV-1 infection as well as inhibiting the syncytia formation are also found identical to those of the parent murine antibody. Moreover, in the presence of human blood mononuclear cells, the chimeric antibody but not BAT123 (mouse IgG1) induces antibody-dependent cellular cytotoxicity. The findings point to the potential usefulness of the chimeric antibody in treating patients infected with HIV-1.
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