Parathyroid hormone-related peptide in the human fetal uro-genital tract.

Using a polyclonal antiserum raised against the first 34 amino acids of human parathyroid hormone-related peptide (PTHrP), we have localized PTHrP throughout the uro-genital tract of the human fetus aged between 8 and 40 weeks. Staining was present in the developing mesonephros, metanephros, gonads and in both the adrenal cortex and medulla. In particular, the developing mesonephric and metanephric renal tubules were intensely positive. Using Northern hybridization analysis we have detected a complex pattern of PTHrP mRNA transcripts ranging in size from 1.4 to 4.5 kb in early second trimester human fetal kidney. The presence of PTHrP in the mesonephros and metanephros provides evidence for a role for PTHrP in the regulation of fetal calcium metabolism. However, its presence in the gonad and adrenal gland invites the possibility of a wider role for PTHrP.

Hemolymph lectin of the pearl oyster, Pinctada fucata martensii: a possible non-self recognition system.

To elucidate a mutual correlation between the hemolymph lectin and hemocytes of the pearl oyster, Pinctada fucata martensii, we searched for common epitopes and ligands. Neither the hemocyte plasma membrane nor cytoplasm was immunoreactive to anti-hemolymph lectin antibody. The hemolymph lectin strongly bound to D-galactose and N-acetyl-D-galactosamine, and the plasma membrane of both granulocytes and agranulocytes had affinity only for D-mannose and N-acetyl-D-glucosamine-binding plant lectins. In the gonad, the hemolymph lectin selectively adsorbed injected horse red blood cells (HRBC), and its hemagglutinating activity probably prevented them from dispersing. Pinctada sp. may possess system of recognition of non-self by the hemolymph lectin.

Immunocytochemical demonstration of the binding of growth-related polypeptide hormones on chick embryonic tissues.

The presence of endogenous growth-related polypeptide hormones, such as growth hormone (GH), somatomadin-C/insulin-like growth factor-1 (SM-C/IGF-1), prolactin (PRL) and Mullerian inhibiting substance (MIS) on chick embryonic tissues have been detected by electron microscopic (EM) immunocytochemistry. Antiserum against GH, anti-SM-C/IGF-1, anti-PRL and anti-MIS were used respectively as primary antibodies for immunolabeling probes by peroxidase (PO) and avidin-biotin complex (ABC)-gold ligands. Cross-reaction studies by ELISA showed negative or weak antigen-antibody interactions. Chick embryos, gonads, and Mullerian ducts (Mds) of various ages were fixed in 2.5% glutaraldehyde for 30 min. Washes in phosphate buffer were administered between each of the following incubations: (i) 2% BSA; (ii) primary antibody; (iii) biotinylated or PO-conjugated secondary antibody; (iv) avidin conjugated with gold particles. SM-C/IGF-1 bindings were negative on 1d embryonic disc, heavily stained on 2d endoderm. However, the GH bindings were found on the embryonic layers of 1d and 2d embryos, and increasing on the luminal epithelial cells of Mds during development. PRL was found in parallel with GH, but in less amount. The 10d Mds were double labeled with anti-SM-C/IGF-1-gold and anti-MIS-PO (MIS-PO), and the results showed SM-C/IGF-I negative, but MIS-PO positive bindings. This study provides the first immunocytochemical evidences for: (i) The presence of GH, SM-C/IGF-1, PRL and MIS bindings on chick embryonic tissues, and further supports their potential role as growth mediators during embryonic development. (ii) The amount of GH and MIS bindings were found correspondingly to their physiological status of Md growth or regression.(ABSTRACT TRUNCATED AT 250 WORDS)

Immunocytochemical localization of inhibin in rat and human reproductive tissues.

Recently, the structures of two forms of inhibin present in human follicular fluid were elucidated from the corresponding cDNA sequences. Using specific antisera generated against the alpha-chain common to both forms, we have examined the cellular localization of inhibin in the male and female rat gonads and in human placental tissue. Specific alpha-inhibin immunoreactivity was localized within the Sertoli cells of a number of tubules in each testes section. However, other adjacent tubules were unstained suggesting a stage-specific production of inhibin. Intense immunostaining was observed in the granulosa cells of ovarian follicles at various stages but not in the thecal cells. Immunostaining was present in the human placenta and limited to the cytotrophoblast cells, suggesting a role of inhibin during pregnancy. The present study demonstrates the probable site of production of inhibin in the gonads and placenta and further implicates this important factor as a key regulator of reproductive functions.

Seasonal variations of lipid fatty acids of boreal freshwater fish species.

1. The fatty acid levels of muscle and liver lipids of perch, vendace and rainbow trout (Salmo gairdneri) cultivated in the same area (for comparison) were monitored. 2. The total lipid content in the muscle of perch (Perca fluviatilis) and vendace (Coregonus albula) was less than 50% of that in rainbow trout and a seasonal variation was clear only in vendace. 3. The relative amounts of polyunsaturated fatty acids as well as omega-3 acids were higher in vendace and perch than in cultivated rainbow trouts. Arachidonic acid content was much higher in vendace and perch than in rainbow trout. The content of monoenes was considerably higher in rainbow trout than in free freshwater fish. 4. The seasonal variations in the degree of unsaturation were small in fish muscle. 5. In the muscle of rainbow trout the relative amount of polyunsaturated fatty acids diminished with the increase of total lipids.

Ovarian function during aflatoxin B1-induced hepatocarcinogenesis in the rat.

Study of the endocrine status, which is often disturbed during hepatocarcinogenesis, is particularly valuable because gonadal function and hormone production regulate hepatic metabolism of the carcinogen. Sex steroids can even promote carcinogenesis. After aflatoxin B1 induction of liver carcinogenesis in adult female Sprague Dawley rats, livers were examined by histology, fluorescence microscopy of the carcinogen and its metabolites, and alphafetoprotein (AFP) assays. Ovarian activity was assessed, and both progesterone and estradiol levels were determined. Administration of a diet containing 10.32% total protein plus 2 ppm aflatoxin B1 was observed to prevent development of liver tumors during the 300 day study period. This finding is especially interesting for the study of populations suffering from malnutrition and exposed to dietary carcinogens. Under study conditions, aflatoxin B1 did not cause elevation of AFP levels, as occurs with other hepatotoxic substances. This absence of a rise in AFP despite liver alterations explains the surprising lack of ovarian modifications. In other experiments, AFP has been shown to cause genital function blockade which leads to reduced levels of hormonal promoters, for example during N2 fluorenylacetamide carcinogenesis. The endocrine reaction implicated in the development of tumors during carcinogenesis thus appears closely related to the nature of the carcinogen, AFP production, and the composition of the diet.

Tissue distribution of amyloid deposits in Abyssinian cats with familial amyloidosis.

The tissue distribution of amyloid deposits was studied in 15 related Abyssinian cats with familial amyloidosis. There was interstitial medullary amyloidosis in the kidneys of all 15 cats but only 11 had detectable glomerular involvement. The thyroid glands, stomach and colon were affected in all cats examined. Most of the cats also had amyloid deposits in the small intestine, spleen, heart, adrenals, pancreas, liver, lymph nodes and bladder. In 50 per cent or fewer of the cats examined, there was involvement of the parathyroids, lung and gonads. The central nervous system was not involved in any of the 3 cats evaluated. In 8 of the cats, no concurrent inflammatory disease could be detected. The tissue distribution of amyloid deposits resembled that found in other breeds of domestic cats with systemic amyloidosis. Despite the wide tissue distribution of amyloid deposits, clinical signs were related to renal amyloidosis. Familial amyloidosis in the Abyssinian cat may represent a valuable spontaneous animal model for the study of Familial Mediterranean Fever in man and the pathogenesis of reactive amyloidosis in general.

Trace metal residues in finfish from Maryland waters, 1978-1979.

Levels of seven heavy metal residues, arsenic, cadmium, chromium, copper, lead, mercury and zinc were monitored in samples of various species of finfish harvested from the Maryland section of the Chesapeake Bay and its tributaries over a two year period (1978-79). Results of the analysis of the edible portions of these finfish are presented along with the species of finfish, date and location of harvest. A number of samples of finfish gonad and liver tissue were analyzed to study the relative level of preconcentration of heavy metals in these tissues compared to the edible (flesh) portion. Results of this study are consistent with other available data for Atlantic Coast finfish. Gonad tissue, when compared to flesh, show enrichment of copper and zinc and decreased mercury and cadmium levels. Liver tissue shows enrichment in copper, zinc and cadmium and generally lower levels of mercury compared to flesh.

Primary structure of histone H2B from gonads of the starfish Asterias rubens. Identification of an N-dimethylproline residue at the amino-terminal.

The complete amino acid sequence (121 residues) of histone H2B from gonads of the starfish Asterias rubens has been established from structural data obtained essentially from large fragments generated by cleavage of histone H2B at aspartyl residues and by limited hydrolysis of the dimer H2A-H2B with mouse submaxillary gland protease. No real sequence homology can be found between the amino-terminal sequence (residues 1-21) of starfish and calf H2B. One non-conservative substitution (serine-32 in calf----lysine-28 in starfish) leads to the presence of a cluster of eight basic residues (sequence 23-30) and to the disappearance of a potential site of phosphorylation. A particular structural feature of starfish histone H2B is the presence of N-dimethylproline at its amino-terminal end. By comparison with N-terminal acetylation, which is commonly found in histones, N-terminal methylation is rarely observed. At the present time the functional significance of the N-terminal methylation as well as that of the proline-rich nature of the amino-terminal sequence of the starfish histone H2B remain to be defined.

Purification and characterization of the gonad lectin of Aplysia depilans.

Extracts of gonads and fertilized eggs of Aplysia depilans contain a D-galacturonic and D-galactose-binding lectin. This lectin reacts strongly with rabbit and human erythrocytes independent of ABO blood groups, weakly with dog, mouse, rat, and chick erythrocytes and not at all or very weakly with sheep erythrocytes. Purification of the gonad lectin was easily achieved, with a high yield, by heating to 70 degrees C, precipitation with ammonium sulfate and affinity chromatography on Sepharose 4B. The purified lectin was found to be a glucoprotein of molecular mass around 55-60 kDa; it stimulates mitogenesis of human peripheral lymphocytes.

Determination of DNA content in the nurse and follicle cells from wild type and mutant Drosophila melanogaster by DNA-Feulgen cytophotometry.

DNA replication patterns in the nurse and follicle cells of wild type and a female sterile mutant, fs(1)1304, of Drosophila melanogaster have been studied by DNA-Feulgen cytophotometry, using a cell dispersal technique that allowed the measurement of DNA amounts in individual nuclei from egg chambers of known developmental stages. DNA-Feulgen values associated with various ovarian nuclei from egg chambers at different stages of development were used to assess a base line DNA content for ovarian tissues and to estimate the extent of DNA replication in the nurse cells and follicle cells of growing and mature egg chambers. Our data show that both the nurse and follicle cells undergo multiple cycles of endonuclear DNA replication and that there may be selective amplification as well as underreplication by portions of the genome in these highly polyploid, ovarian cells. Alternative models are proposed to account for the DNA replication patterns observed. Comparisons of DNA-Feulgen levels in wild type ovarian nuclei with those found for the fs(1)1304 mutant and its heterozygote in the balanced stock fs/FM3, show that equivalent DNA levels are present in follicle cell nuclei from all three types of females. Nurse cell nuclei in the homozygous fs stock, however, fail to achieve the same high DNA levels observed in both fs/FM3 and wild type nurse cell nuclei. Although the nuclei of follicle cells in ovaries from fs/fs females appear morphologically like those surrounding egg chambers in wild type ovaries, nurse cell nuclei from mutant females show a more compacted organization of their chromatin than found for nurse cell nuclei from wild type ovaries at similar developmental stages. Our findings suggest that a major effect of the fs(1)1304 mutation may be on the coiling behavior of chromatin and the conformation of DNA-protein moieties in both nurse cell and follicle cell nuclei. These changes in chromatin structure apparently are manifest by perturbations in DNA replication patterns and normal gene function in these biosynthetically active cells.

Effect of tamoxifen on H-Y antigen expression and gonadal development in chicken embryos.

Avian species follow the ZW/ZZ system of sex determination, which the female is heterogametic and expresses H-Y (or, more appropriately, 'H-W') antigen. We present the results of an investigation into the effects of the antiestrogen, tamoxifen, on gonadal differentiation and H-Y antigen expression in chickens. When given at doses of 0.25-2 mg per egg immediately before incubation, tamoxifen blocked regression of the right gonad in a significant number of 14-day-old female embryos. The nonregressed right gonad had a testis-like external appearance and, in some cases, contained what appeared to be spermatogenic tubules. Tamoxifen had no histologically detectable effect on the differentiation of the left ovary or the testes. In spite of tamoxifen's histological effects on right female gonads, it did not masculinize the steroidogenic capabilities of these gonads. Whether obtained from drug- or vehicle-treated embryos, the left and right female gonads always contained appreciable amounts of estrogen. In contrast, testes obtained from either drug- or vehicle-treated embryos did not contain detectable amounts of estrogen. Tamoxifen reduced the H-Y antigen levels in female liver and gonads. In both left and right female gonads, the reduction was to male levels. In female livers, tamoxifen reduced H-Y antigen to levels intermediate between those of normal males and females. Thus, the expression of H-Y antigen in both gonadal and nongonadal tissue is estrogen dependent, but the dependency appears to be more stringent for gonadal tissue.(ABSTRACT TRUNCATED AT 250 WORDS)

Variability in the apparent molecular weight of inhibin.

An in vitro bioassay based on suppression of GnRH-stimulated FSH secretion by pituitary cells in culture was used to monitor inhibin activity after dialysis, gel filtration or polyacrylamide gel electrophoresis of protein preparations from a variety of gonadal secretions and extracts under native and dissociating conditions. The suggestion that inhibin is a peptide of molecular weight less than 5000 was not confirmed. Although some fractions of low molecular weight suppressed FSH secretion, the amount of activity was low and the dose response curves were not parallel with a standard preparation of inhibin. Under most conditions, inhibin eluted with an apparent molecular weight of about 90 000. However, gel filtration of rete testis fluid protein in 1 M acetic acid resulted in elution of inhibin activity with a lower apparent molecular weight and with polyacrylamide gel electrophoresis in 0.1% (w/v) sodium dodecylsulfate, the apparent molecular weight was 30 000. It is concluded that inhibin is a protein which tends to aggregate and coelute with larger molecules.

Chromosomes and sex differentiation in eutherians.

In order to learn more about the role of sex chromosome-dependent gene products in gonadogenesis, changes in protein patterns were studied during gonadal development. Two-dimensional gel electrophoresis analysis revealed specific proteins in both sexes at all developmental stages. Evidently the gonads are not indifferent by biochemical criteria at any developmental stage and express several specific genes from the onset of differentiation. To correlate these polypeptides with the sex chromosomes, proteins were investigated in human-rodent somatic cell hybrids and in genetically identical cell clones differing in one sex chromosome only. On two-dimensional gels one Y-dependent polypeptide was found with similar characteristics (relative molecular mass and isoelectric point) as an early testicular polypeptide. Its identity, however, remains to be proven.

A morphological, enzyme-cytochemical, and physiological study of the blood-gonad barrier in the hermaphroditic snail Lymnaea stagnalis.

In the hermaphroditic pulmonate snail Lymnaea stagnalis a blood-gonad (blood-testis) barrier appears to exist. Septate junctions between Sertoli cells and epithelial cells of the neck areas of the gonadal acini constitute this barrier; they separate the male from the female compartment. Experiments with tracer substances (colloidal gold particles, lanthanum nitrate, tannic acid) showed that the basal lamina around the acini hardly forms a barrier; only the larger colloidal gold particles do not pass this lamina. Physiological, the blood-gonad barrier is apparent in studies on the composition of gonadal fluid, which differs considerably from that of haemolymph. The osmolarity and the concentration of protein and amino acids in gonadal fluid exceed those of haemolymph. As to the major ions, in the gonadal fluid Na+ is partly replaced by K+, and HCO-3 is almost totally replaced by Cl-. Such a distribution of HCO-3 and Cl- is indicative of metabolic acidosis. The cytochemical localization of carbonic anhydrase activity in cells lining the acinar lumen (Sertoli cells, epithelial cells) suggests that these cells are involved in the process of ion exchange. The metabolic acidosis in the gonad might result from the anaerobic production of lactate and succinate by Sertoli cells; these cells lack the enzymes cytochrome oxidase, lactate dehydrogenase, and succinate dehydrogenase. Spermatogenic cells, on the other hand, do possess these enzymes. This probably indicates that these cells metabolize lactate and succinate secreted by Sertoli cells.