Anti-luteinizing Hormone Research Articles

HLA-B*0702 class-I allele, anti-FSH, anti-LH, and vitamin D3: Potential links with polycystic ovary syndrome in women of Erbil city, Iraq

Polycystic Ovary Syndrome (PCOS) is a widespread endocrine-reproductive-metabolic condition with severe implications for females’ health. The role of four essential parameters on PCOS, including Human Leukocyte Antigens (HLA) represented by the HLA-B*0702 allele, anti-Follicle Stimulating Hormone (anti-FSH) antibodies, anti-Luteinizing Hormone (anti-LH) antibodies, and vitamin D3 was investigated. A total of 100 samples were collected from Kurdish women attended the Maternity Teaching Hospital and some private hospitals/laboratories in Erbil City from October, 2021 to January, 2022. The samples were genotyped using a PCR-based technique with specific sequence primers. The levels of anti-FSH and anti-LH antibodies were determined using an Enzyme-Linked Immunosorbent Assay (ELISA), while vitamin D3 levels were measured by an Electrochemiluminescence (ECL) test on Cobas e411 immunoassay system. The Odds Ratio (OR) of 2.167 at a 95% Confidence Interval (CI) of 0.8167 to 6.330, indicated an essential link between the HLA-B*0702 allele and a risk of PCOS. Anti-FSH and anti-LH antibodies were significantly greater in PCOS patients, notably infertile women, than in healthy controls. A significant positive linear correlation was observed between antibodies against FSH and LH in patients. Most patients and controls had hypovitaminosis D3, with a significant difference (P-value  0.01). The results indicated that the HLA-B*0702 allele is associated with PCOS susceptibility and could be used as an immunogenetic marker. It also supported the idea that anti-FSH and anti-LH antibodies are naturally presence antibodies in PCOS patients instead of signs for autoimmunity. Women suffer from PCOS are also more prone to develop vitamin D3 deficiency.

Flow immunosensing system utilizing antibodies immobilized on beads coated with poly-2-[3-(methacryloylamino)propylammonio]ethyl 3-aminopropyl phosphate

In order to increase the signal-to-noise ratio of immunosensors, a flow immunosensing system was fabricated containing poly-2-[3-(methacryloylamino)propylammonio]ethyl 3-aminopropyl phosphate ( poly-3MAm3AP) at the antigen–antibody reacted interface. Poly-3MAm3AP is a polymer containing phosphorylcholine groups (which act to attract water) and amino groups (useful for the covalent attachment of receptors). Anti-luteinizing hormone (LH) antibody-immobilized beads coated with poly-3MAm3AP were prepared and packed into the reactor column of a flow immunosensing system. Nonspecific anti-hemoglobin antibodies were also used in order to estimate the level of nonspecific adsorption. The fabricated immunosensing system not only significantly reduced nonspecific adsorption, but also detected signal intensity with superior signal-to-noise, thus demonstrating the potential of this approach for the commercialization of immunosensing technologies.

Roles of Basal Levels of Circulating Luteinizing Hormone on Follicular Maturation in the Pregnant Golden Hamster (Mesocricetus auratus).

To investigate physiological roles of basal levels of circulating LH on follicular development during the second half of pregnancy in golden hamsters, we treated animals with anti-luteinizing hormone (A/LH) or normal horse serum (control serum) at 1100 h on day 12 of pregnancy (day 0 of pregnancy=sperm positive in vaginal smear). Thereafter, changes in plasma concentrations of immunoreactive (ir)-inhibin, estradiol-17β, progesterone and testosterone were determined until 48 h after the treatment; ovaries were also examined immunohistochemically. Furthermore, pregnant golden hamsters were injected with human chorionic gonadotropin (hCG) to determine the effects of immunoneutralization of LH on follicular activity. Plasma concentrations of ir-inhibin and estradiol-17β decreased significantly within 24 h after A/LH treatment, while plasma concentrations of progesterone and testosterone were unchanged. The number of follicles ovulating in response to hCG in the A/LH treated groups (7.0 ± 3.4, n=3) was significantly lower than in the control serum treated group (34.6 ± 4.8, n=5) at 24 h after the treatment. Immunostaining of inhibin α subunit was observed in granulosa cells of healthy and atretic large follicles at 24 h after A/LH treatment, whereas immunopositive reaction of aromatase was not observed in atretic follicles. Positive immunoreaction of 17α-hydroxylase/C17, 20-lyase (CYP17) was observed in theca cells of healthy follicles but not in atretic follicles. These results clearly demonstrate that basal levels of circulating LH are important for maturation of antral follicles and secretion of ir-inhibin and estradiol-17β, but not for progesterone during the second half of pregnancy in the golden hamster.

Steroid 5α-reductase type 1 immunolocalized in the anterior pituitary of intact and castrated male rats

The localization of 5 alpha-reductase was immunohistochemically studied in the anterior pituitary of male rats, using a polyclonal antibody against 5 alpha-reductase rat type 1. The immunoreactive cells were concentrated in the central region and on the border of the intermediate lobe in the anterior pituitary, but not in the intermediate or posterior lobe. The immunoreaction was located mostly in the cytoplasm and occasionally in the cell nuclei. The immunoreactive cells showed alterations in size and number and in the intensity of the immunoreaction after gonadectomy. One week after castration, the cells became larger and the immunoreactivity increased. Two weeks after castration, the number of immunoreactive cells increased. Double immunostaining using anti-luteinizing hormone beta-subunit or anti-follicle stimulating hormone beta-subunit antibody revealed that most of the cells containing 5 alpha-reductase were gonadotrophs. Electron microscopically, the immunoreactive cells showed lamelliform rough endoplasmic reticulum and a depletion of secretory granules 1 week after castration. One week later, the rough endoplasmic reticulum was developed and dilated and the number of secretory granules increased. These results suggest that 5 alpha-reductase is located in the gonadotrophs of rat anterior pituitary and that it is involved in the feedback regulation of gonadotropin secretion by androgens.

Detection of epitopes on follicle-stimulating hormone and FSH-antiserum-induced suppression of bioactivity of follicle-stimulating hormone and luteinizing hormone

There are currently two major approaches to hormonal male contraception. One relies on testosterone (analogs) either alone or in combination with gonadotropin releasing hormone (GnRH) (analogs or immunizations), the other on immunizations against follicle-stimulating hormone (FSH). Theoretically, the latter method will suppress spermatogenesis whilst not interfering with libido. An absolute requirement is, however, that an anti-FSH vaccine does not induce anti-luteinizing hormone (LH) antibodies (LH being responsible for the induction of testosterone which is necessary-to maintain libido). In this report we show that when whole FSH is used for vaccination, in most cases in addition to biological activity against FSH, anti-LH activity is also induced. By systematic analysis of the antisera raised with FSH using systematic epitope scanning (PEPSCAN) we found differences between the FSH-specific and FSH-nonspecific sera. Only the FSH-specific antiserum contained antibodies that recognized amino acid sequence 37–55 on the β-subunit in a linear manner. Because antibodies against this epitope have not been found in the cross-reactive sera this epitope forms a prime candidate for an anti-FSH contraceptive vaccine

Characterization by means of Delaunay triangulation and Voronoi paving of the influence of anti-hormone and/or anti-growth factor antibodies on the in vitro cell growth of human colorectal neoplastic cell lines

A new tool is described which makes it possible to evaluate directly the influence of various growth factors on in vitro neoplastic cell growth on the one hand and to look at a concept of differentiation in terms of population dynamics, on the other. This tool relies upon the digital cell image analyses of Feulgen-stained nuclei and the mathematical method of Voronoi paving. This technique enabled us to characterize the influence on the proliferation and the differentiation of the HCT-15 and LoVo colorectal cell lines of anti-gastrin (G), anti-estradiol (E(2)), anti-epidermal growth factor (EGF), anti-luteinizing hormone-releasing hormone (LHRH), and anti-transforming growth factor alpha (TGF alpha) and beta (TGF beta) antibodies. Two variants were set up with respect to each of the two cell lines, i.e, one growing in culture medium supplemented with 5% fetal calf serum (FCS) and another supplemented with 1% FCS+10 nM G+10 nM E(2). The data show that it is possible to characterize the cell clone structure and to assess growth rate concomitantly by direct cell counts. It further appears that while the anti-hormone and/or anti-growth factor antibody-induced effects on growth were relatively similar, these effects were in sharp contrast at the level of cell clone architecture.

Effects of Injection of Anti-Luteinizing Hormone (LH)-Releasing Hormone Serum and Anti-Gonadotropin-Releasing Hormone-Associated Peptide Serum into Neonatal Rats on LH and Follicle-Stimulating Hormone Cells1

Relatively little is known regarding the potential importance of LHRH and of gonadotropin-releasing hormone-associated peptide (GAP) on the postnatal development of gonadotrophs. We investigated the effects of administration of anti (A)-LHRH serum or A-GAP serum to neonatal rats on the development of LH and FSH immunoreactivity in anterior pituitary gland (APG) cells. Serum (sheep non-immune [NSS], sheep A-LHRH, sheep A-LHRH/GAP [which bound LHRH and GAP], rabbit non-immune [NRS], or rabbit A-GAP) was injected s.c. into neonatal female and male rats on Days 1 and 3 or Days 1, 3, 5, and 7 after birth. Pups were killed on Day 5 or 9, two days after the last injection. The percentages of APG cells immunoreactive for LH or FSH increased from Day 1 to Day 5 and did not change between Days 5 and 9 in female pups treated with NSS or NRS. There was a trend for the percentages of LH and FSH cells to increase from Day 1 to Days 5 and 9 in male pups treated with NSS or NRS, but the increases were not statistically significant. In both females and males, treatment with antisera that recognized LHRH reduced the percentage of FSH cells on Day 5 and the percentages of LH and FSH cells on Day 9. Treatment with A-GAP was without effect in both sexes. There were similar percentages of LH and FSH cells in females and a lower percentage of FSH than of LH cells in males in the Day 5 and 9 controls.(ABSTRACT TRUNCATED AT 250 WORDS)

Prostatic hypoplasia in bonnet monkeys following active immunization with semisynthetic anti-LHRH vaccine

An active immunization study using semisynthetic anti-luteinizing hormone releasing hormone (LHRH) vaccine was undertaken in adult male bonnet monkeys ( Macaca radiata). Four male bonnet monkeys were immunized with LHRH-DT vaccine (100 μg/monkey). When the elevation of serum anti-LHRH titers reached an effective concentration, a remarkable decrease in serum testosterone level was observed. The sperm counts at 12 and 20 weeks following immunization in all immunized monkeys were reduced markedly. One representative animal with high antibody titer along with an age-matched control animal was sacrificed 20 weeks following primary immunization. The prostate weight of the immunized monkey was reduced to one-third (0.027) of the control (0.085). A reduction was also observed in other reproductive organs. Remaining animals are being studied for a reversible effect of immunization. Thus the study demonstrates that the size of the prostate can be reduced significantly by inhibition of hypothalamic-pituitary-testicular axis using this anti-LHRH vaccine.

Purification of gonadotropes and intracellular free calcium oscillation. Effects of gonadotropin-releasing hormone and interleukin 6.

The intracellular free calcium concentration ([Ca2+]i) in single gonadotropes was measured with a calcium-sensitive fluorescent dye indo-1 or fura-2 and a digital imaging fluorescence microscopic system to determine how interleukin-6 (IL-6) increases release of gonadotropins. IL-6 induced an increase in the basal [Ca2+]i or the amplitude of spontaneous oscillation of [Ca2+]i in gonadotropes in a mixed population. Gonadotropin-releasing hormone (Gn-RH) induced a biphasic increase in [Ca2+]i, a transient increase, and then a prolonged increase. These effects were inhibited by the absence of extracellular calcium or pretreatment with calcium channel blockers, cobalt or nifedipine. Next, purified gonadotropes were prepared by fluorescence-activated cell sorting and argon laser treatment of the cells. Gonadotropes labeled with anti-luteinizing hormone antibody were sorted by fluorescence-activated cell sorting and then cultured as monolayers for 24-48 h. In this way, gonadotropes were concentrated from 5-10% to 70-85% from whole pituitary cells. After relabeling with anti-luteinizing hormone antibody, 100% purified gonadotropes were obtained by killing other types of cells with an argon laser. Gonadotropin-releasing hormone induced almost the same responses of [Ca2+]i in the purified cell population as in the mixed cell population, but IL-6 did not affect [Ca2+]i in the purified gonadotropes. These results suggest that IL-6 affects calcium mobilization in gonadotropes indirectly via paracrine pathways.

Pituitary adenomas: immunohistochemical study of 167 cases

One hundred and sixty seven cases of pituitary adenoma were analysed using the immunocytochemical method of the Avidin-Biotin Complex (ABC), described by Hsu et al. (1981). Six pituitary anti-hormones were utilized: anti-prolactin (aPRL) at a 1:1,500 dilution; anti-growth hormone (aHGH) at a 1:4,000 dilution: anti-adrenocorticotrophic hormone (aACTH) at a 1:3,000 dilution; anti-thyrothrophic hormone (aTSH) at a 1:3,000 dilution; anti-luteinizing hormone (aLH) at a 1:1,000 dilution; and a anti-follicle-stimulating hormone (aFSH) at a 1:300 dilution. Incubation period was 14 to 16 hours at 4 degrees C. The survey of clinical, laboratory and radiological data of cases of pituitary adenomas was performed after reading the stained slides using the immunocytochemical method. Of the 167 cases of pituitary adenomas, 136 (81.4%) disclosed a positive immunoreaction to one or more anti-hormones, and the positivity index of neoplastic cells varied from 1 to 90%. The immunoreaction was positive exclusively to one anti-hormone in 80 cases (58.8%) and to two or more anti-hormones in 56 cases, and the association most frequently found was between both aPRL and aHGH. The positivity to the immunoreaction was distributed as follows: -100 cases were positive for aPRL, exclusively in 4 cases; -65 cases were positive for aHGH, exclusively in 22 cases; -31 cases were positive for aACTH, exclusively in 8 cases; -5 cases were positive for aTSH, exclusively in one case; -one patient presented an adenoma positive to aLH and another patient to aFSH.

Preclinical safety studies with anti-luteinizing hormone releasing hormone (LHRH) vaccine

Previous studies with a semi-synthetic vaccine indicated the utility of immunization against luteinising-hormone-releasing-hormone (LHRH) in prostate cancers. To overcome the limitations of the previous vaccine, which caused carrier induced suppression of antibody reponse on repeated immunizations and was costly to synthesize, two recombinant vaccines were designed, in which diptheria or tetanus toxoid used as carriers were replaced by 4–5 T non B peptides. The paper reports the immunogenecity, efficacy and safety of these multimer vaccines in rats, a homologous experimental animal. All animals generated anti-LHRH antibodies, which caused the decline of testosterone to castration levels at and above 0.15 OD units of antibody titres. The prostate was significantly atrophied in all animals immunized with these vaccines.

The effect of luteinizing hormone releasing hormone and anti-luteinizing hormone releasing hormone antibodies on chorionic gonadotropin and progesterone secretion by human placental villiin vitro

Gonadotropin releasing hormone has been located and found to be secreted by the human placenta in culture. Addition of the releasing hormone upto 1μg concentration in the placental cultures brings about stimulation of chorionic gonadotropin and progesterone secretion. Higher amounts of the decapeptide has an inhibitory influence on both the gonadotropin and the steroid production. The action of the releasing hormone on the placenta could be blocked by the anti-luteinizing hormone releasing hormone monoclonal antibodies indicating a possible site of action of the antibodies for control of fertility

Preparation of anti-human chorionic gonadotropin and anti-luteinizing hormone vaccine using oligopeptides specific to beta-subunit of LH and HCG; contraceptive

Preparation of anti-human chorionic gonadotropin and anti-luteinizing hormone vaccine using oligopeptides specific to beta-subunit of LH and HCG; contraceptive

Extra-pituitary action of gonadotropin releasing hormone: possible application

Chronic administration of a potent gonadotropin releasing hormone inhibits ovulation in women. The suppression of gonadal function during long term treatment with the GnRH analogues is ascribable to inhibition of gonadotropin secretion caused by the down regulatory action of the decapeptide at the pituitary level. Reduced progesterone production with premature onset of menstruation has been observed in women injected with the agonist during the midluteal phase. The decapeptide however, has no effect on in vitro human ovarian steroidogenesis. Specific receptors for GnRH have been located on rodent ovarian cells, but corpora lutea of rhesus monkey and human ovaries seem to lack these receptors. The luteolytic effect in women thus appears to be central in origin and not a direct effect on the corpus luteum. Recently, a superactive agonist of GnRH given around the peri-implantation period has been shown to terminate pregnancy in baboons. Monoclonal antibodies against GnRH administered during the same period in a fertile cycle also abrogated pregnancy in these animals. Using immuno-enzymatic techniques GnRH has been localized on the placenta. GnRH also exerts a stimulatory effect on hCG production by the placental villi maintained in culture. Addition of anti-luteinizing hormone releasing hormone antibodies blocks this effect completely. It seems that placenta is the only other tissue besides the pituitary where GnRH has probably a regulatory role in the human female.

Study of the direct effect of LHRH agonist on testicular 17-hydroxylase and 5 α-reductase activities in non-hypophysectomized adult rats treated with an anti-luteinizing hormone serum

In order to study both direct and pituitary-mediated mechanisms of action of the LHRH analogue [D-Ser(TBU) 6, des-Gly-NH 2 10] LHRH ethylamide upon testicular steroidogenesis in adult rat, we compared the effects of the agonist when administered alone or concomitantly with an anti-LH serum to non-hypophysectomized rats. Testicular steroid contents and in vitro progesterone and testosterone metabolism were determined. Anti-LH serum administration was able to prevent 5 α-reductase stimulation by the agonistic peptide, but not the inhibition of 17-hydroxylase activity. These data suggest that modulation of 17-hydroxylase involves both direct and pituitary-mediated processes, while 5 α-reductase stimulation is mainly if not only due to a pituitary-mediated mechanism.

The role of endogenous opioids in the blockade of reproductive function in the rat following exposure to acute stress

The present experiments investigated the involvement of endogenous opioids in the preovulatory disruption of luteinizing hormone release and ovulation following exposure to an acute stressor. The effect of exposure to an acute stressor and/or treatment with naloxone hydrochloride on the proestrus/estrus display of lordosis behavior is also reported. Electric shocks (2 mA) delivered at 10-min intervals for three hours during the period of the preovulatory luteinizing hormone surge in adult cyclic female rats resulted in the blockage of luteinizing hormone release, decreased lordosis behavior, and inhibition of ovulation. The anti-luteinizing hormone and anti-ovulatory actions were antagonized by the administration of naloxone hydrochloride shortly prior to the application of the stressor, which suggests that the observed blockage of ovulation is mediated by endogenous opioids known to be elevated in response to a stressor. It is argued that the most likely mechanism by which endogenous opioids inhibit luteinizing hormone release and ovulation is by inhibiting luteinizing hormone releasing hormone release, and/or decreasing its production in the hypothalamus. In contrast, naloxone only partially antagonizes the anti-lordotic actions of electric shock. It is therefore further argued that the inhibition of luteinizing hormone releasing hormone release alone is not sufficient to explain the dramatic decrease in lordosis behavior observed following exposure to an acute stressor. Adrenal progesterone released in response to adrenocorticotropic hormone release following exposure to an acute stressor, in association with inhibited luteinizing hormone releasing hormone release is hypothesized to be responsible for the observed decrease in lordotic behavior, and the inhibition of luteinizing hormone release.

Blockade of post-partum ovulation by anti-luteinizing hormone serum in lactating rats.

To examine the suppressive effect of suckling stimulus and ovarian progesterone and their interaction on the resumption of ovarian cyclicity during lactation, peripheral plasma progesterone levels and the length of diestrous periods were studied by using lactating rats of which post-partum ovulation was blocked by anti-LH serum. Plasma progesterone levels were less than 5.6ng/ml in the rats treated with antiserum despite of their size of litter that had been nursed. The diestrous period in antiserum treated rats nursing 12 pups (15.4±1.4 days) was shorter than that in the intact rats nursing the same number of pups (26.2±1.1 days). In the rats nursing 2 pups, antiserum treatment also shortened the di-estrous period (6.5±0.1 days) and this value was the same as those in the intact or anti-serum treated rat of which pups were removed immediately after parturition. Plasma progesterone levels before and after removal of pups were studied in antiserum treated rats nursing 12 pups up to 10 days after parturition. They were received progester-one injection from 3 to 10 or to 16 days after parturition and were deprived of their litter 10 days after parturition. When progesterone was injected from 3 to 16 days after parturi-tion, the diestrous period following removal of pups was prolonged to 8.7 days. Cessation of progesterone treatment 10 days after parturition markedly shortened the diestrous period following removal of pups (3.4 days) and reduced the plasma progesterone to the basal level on the day following removal of pups.

Immunohistochemical localization of renin in luteinizing hormone-producing cells of rat pituitary.

The location of renin (EC 3.4.99.19) in rat pituitary was determined by the peroxidase-antiperoxidase immunohistochemical technique. By using antisera prepared with purified rat renal renin, an immunoreactive substance was localized within ovoid cells scattered throughout the anterior pituitary. These cells were shown to be luteinizing hormone-producing cells by staining with anti-luteinizing hormone antisera in adjacent sections. By using the double staining method, the renin-containing cells were differentiated from cells containing corticotropin, thyrotropin, growth hormone (somatotropin), and prolactin (mammotropin). These results suggest a possible local role for renin in the anterior pituitary.

Immunoperoxidase staining of primate pituitaries with antibodies against the beta subunits of human pituitary glycoprotein hormones.

The immunocytochemical specificity of antibodies against the beta subunits of human pituitary glycoprotein hormones was tested on human and monkey hypophyses. Anti-luteinizing hormone (LH) beta stained only gonadotrophic cells in both species, and anti-thyroid-stimulating hormone (TSH) beta only thyrotrophic cells. In monkeys, anti-follicle-stimulating hormone (FSH) beta demonstrated only gonadotrophs, but in man the antibody stained thyrotrophs equally well. Staining of the two cell types was diminished to a similar degree by dilution of the anti-FSH beta antibody. Human gonadotroph heterogeneity was revealed by absorption of anti-FSH beta with LH beta or TSH beta, which abolished immunostaining of a subset of gonadotrophs. Limitations in the immunocytochemical specificity of antibodies against beta subunits of pituitary glycoprotein hormones may be a consequence of incomplete purity of the generating antigens, structural overlap among the beta subunit of the three hormones, or a combination of these.

Alteration of the serum follicle-stimulating hormone to luteinizing hormone ratio in the cyclic hamster treated with antiluteinizing hormone: relationship to serum estradiol, free antiluteinizing hormone, and superovulation.

The serum concentrations of unbound LH, FSH, estradiol (E2), and free antibovine LH serum (anti-LH) were determined by RIA in hamsters destined to superovulate (∼32 ova) after a single sc injection of 0.1 ml equine anti-LH at 0900 h on day 1 of the cycle (morning of ovulation). As a control, another group of hamsters was injected with normal horse serum (NHS) at 0900 h on day 1. Animals injected with anti-LH or NHS were decapitated at various times over the next 6 days. Interfering LH antibodies were removed by column chromatography in order to determine the serum concentrations of free FSH and LH by RIA. Serum concentrations of unbound FSH were similar throughout the cycle in animals treated with anti-LH or NHS. On day 2 of the cycle (24 h after anti-LH), free LH levels were less than 6 ng/ml (the lower limit of sensitivity of the RIA on the volume of serum assayed), free anti-LH titers were extremely high, and E2 levels were less than 10 pg/ml. The absence of detectable unbound LH and normal FSH in sera...