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Biomedical subjects

E C Weir

Publications and source records attributed to E C Weir.

At least 37 records · Page 2Linked to original sources
PubMed1992

Abundant expression of parathyroid hormone-related protein in human amnion and its association with labor.

In animal models, parathyroid hormone-related protein (PTHrP) increases placental calcium transport and inhibits contraction of uterine smooth muscle. The present studies were undertaken to characterize the expression of PTHrP in human uteroplacental tissues. PTHrP mRNA was identified by Northern analysis as a single species (approximately 1.8 kilobases) in human amnion, chorion, placenta, decidua, and myometrium. The most abundant signal was seen in amnion, where it was 10-400 times that in the other uteroplacental tissues. PTHrP mRNA abundance was decreased in amnion (but not in the other tissues) following the onset of labor (P less than 0.001). PTHrP mRNA in amnion appeared to be translated to a bioactive peptide, as PTHrP bioactivity and immunoreactive PTHrP in amnion correlated closely with PTHrP mRNA content (r = 0.86 and 0.95, respectively; P less than 0.05 and P less than 0.01). Amniotic fluid contained PTHrP, 21 +/- 6 pmol/liter (n = 10) at 16 weeks and 41 +/- 9 pmol/liter (n = 7) at 38 weeks (P = 0.05). These concentrations equaled or exceeded those found in plasma of patients with hypercalcemia secondary to PTHrP. After rupture of the fetal membranes, PTHrP mRNA in amnion was decreased by 78% (P less than 0.0001). This decrease appeared to be specific for PTHrP mRNA, as glyceraldehyde-3-phosphate dehydrogenase mRNA was unchanged following rupture of membranes. Like PTHrP mRNA, PTHrP bioactivity and immunoreactive PTHrP in amnion decreased significantly following rupture of membranes (P less than 0.03 and P less than 0.01, respectively). Since PTHrP is a potent antagonist of uterine muscle contraction, the decrease of PTHrP following rupture of the fetal membranes may play a key role in the onset of labor.

Amnion↗
PubMed1992

Synthetic parathyroid hormone-like protein (1-74) is anabolic for bone in vivo.

Parathyroid hormone-related protein (PTHRP) has recently been purified from human tumors associated with the syndrome of humoral hypercalcemia of malignancy. The gene encoding PTHRP has been cloned, and based on predicted amino acid sequence, polypeptides comprising the first 36 [36Tyr(1-36) PTHRP amide] and 74 [(1-74)PTHRP] amino acids have been synthesized. Human (h) PTHRP (1-36) and (1-74) are potent bone-resorbing agents, and are catabolic for bone in vivo when given continuously at high doses. Bovine parathyroid hormone (bPTH) (1-34) is also catabolic for bone at high dose levels, but when given in low doses for weeks to months, it is anabolic. Although PTHRP possess several PTH-like properties in bone, hPTHRP (1-34) is reported to be only weakly anabolic in vivo. As polypeptide length influences PTHRP action, we evaluated hPTHRP(1-74) as an anabolic agent for bone in vivo. Twenty-four 4-week-old male Sprague-Dawley rats were given daily subcutaneous injections of hPTHRP(1-74) (1 and 2 nmol/100 g body weight, bw), bPTH(1-34) (4 nmol/100 g bw) or vehicle. Rats were sacrificed on day 12, and serum calcium, phosphorus, and 1,25 dihydroxyvitamin D and femoral bone dry weight, calcium content, and hydroxyproline content were measured. Serum calcium and phosphorus were equivalent in all groups. A significant increase in dry bone weight was observed in both PTHRP-treated groups compared with controls. PTHRP also caused a significant, dose-dependent increase in bone calcium and hydroxyproline content. Results of these studies indicate that PTHRP (1-74) is anabolic for bone in vivo when administered at low-dosage levels for a prolonged period.

Animals↗
PubMed1992

Endometrial venous aneurysms in three New Zealand white rabbits.

Hematuria in rabbits has been associated with uterine adenocarcinoma, uterine polyps, renal infarction, urolithiasis, cystitis, bladder polyps, and pyelonephritis. Three adult female New Zealand White rabbits (Oryctolagus cuniculus) developed apparent hematuria, as suggested by blood in their excreta pans. They had been immunized with antigen-adjuvant emulsions, but had uneventful clinical histories. Physical examination disclosed no abnormalities, and laboratory tests, including hematology, serum chemistries, urinalyses, urine cultures, ultrasonography, and intravenous pyelography disclosed mild anemia, hematuria, and proteinuria in two of the rabbits. Antibiotic therapy failed to alleviate clinical signs. Two rabbits were euthanized because of persistent urogenital bleeding and the third rabbit underwent exploratory laparotomy and ovariohysterectomy. Multiple endometrial venous aneurysms were present in the uteri of all rabbits and urogenital bleeding was attributed to episodic bleeding from these lesions. Varices and aneurysms of uterine subserosal and myometrial venous plexuses, but not of endometrial vessels in women have been reported. To our knowledge, endometrial venous aneurysms have not been reported in animals previously. Our findings indicate that the differential diagnoses for sporadic apparent hematuria in female rabbits should include endometrial aneurysms.

Aneurysm↗
PubMed1990

Regulation of parathyroid hormone-related peptide gene expression by cycloheximide.

A novel parathyroid hormone-related peptide (PTHRP) has been isolated from tumors associated with the syndrome of humoral hypercalcemia of malignancy. The human PTHRP gene appears to use multiple promoters and contains alternatively spliced 3' exons which give rise to three PTHRP mRNA classes, each bearing multiple copies of an AU motif that has been associated with mRNA instability. We report here that inhibition of protein synthesis leads to the super-induction of PTHRP mRNA expression in a number of human and rat cell lines. This phenomenon was found to reflect both an increase in the rate of PTHRP gene transcription and a stabilization of PTHRP mRNAs. The transcriptional mechanism appears to preferentially involve the activity of a short downstream promoter of the gene, which is presumed to be regulated by a labile repressor protein. Our findings indicate that both transcriptional and posttranscriptional mechanisms may be important control points in the regulation of PTHRP expression in normal and malignant cells.

Actins↗
PubMed1990

Parathyroid hormone-related peptide gene is expressed in the mammalian central nervous system.

A parathyroid hormone-related peptide (PTHRP) has been identified in human tumors associated with the syndrome of humoral hypercalcemia of malignancy. While parathyroid hormone (PTH) gene expression appears to be limited to the parathyroid glands, PTHRP mRNA has been identified in a variety of normal tissues. To investigate the apparent expression of the PTHRP in the central nervous system, we examined extracts of whole rat brain for PTHRP bioactivity by measuring adenylate cyclase-stimulating activity (ACSA) in a PTH-sensitive assay. Extracts consistently contained ACSA and this activity was completely inhibited by a PTHRP antiserum but was unaffected by a PTH antiserum. ACSA was found in a number of anatomic subregions of rat brain, being greatest in the cortex and telencephalon. RNase protection analysis revealed PTHRP transcripts in total RNA prepared from whole rat brain and from the same anatomic subregions. By in situ hybridization histochemistry, we found that the highest levels of PTHRP gene expression occurred in neurons of the cerebral cortex, hippocampus, and cerebellar cortex. These studies demonstrate that both PTHRP mRNA and biological activity are present in a number of regions of rat brain. The widespread expression of this peptide by multiple types of neurons suggests that the PTHRP may play a general role in neuronal physiology.

Adenylyl Cyclases↗
PubMed1990

Intrauterine occupancy controls expression of the parathyroid hormone-related peptide gene in preterm rat myometrium.

The parathyroid hormone (PTH) and parathyroid hormone-related peptide (PTHRP) genes are members of a gene family. Whereas PTH is a classical peptide hormone, mounting evidence suggests that the PTHRP may have predominately local actions. We report here that the PTHRP gene is expressed in rat myometrium, with a major peak in PTHRP mRNA expression occurring in the 48 hr immediately preceding parturition. A similar peak in peptide content was found in tissue extracts by biological and immunological assays, but the PTHRP could not be detected in the peripheral circulation or in uterine vein plasma during late gestation. By in situ hybridization histochemistry, PTHRP mRNA was demonstrated in both the longitudinal and circular layers of smooth muscle but was absent in the endometrium. The rise in myometrial PTHRP mRNA in late gestation was dependent upon intrauterine occupancy; it was greatly reduced or absent in nongravid uterine horns. These findings indicate that the expression of the PTHRP gene in preterm myometrium is under the control of a local stimulus and suggest that the PTHRP may play a paracrine or autocrine role in the uterus during the antepartum period, possibly involving myometrial contractility.

Animals↗
PubMed1990

The parathyroid hormone-related protein stimulates human osteoblast-like cells to secrete a 9,000 dalton bone-resorbing protein.

The mechanism by which parathyroid hormone-related protein (PTH-RP) stimulates bone resorption is not known. Like certain other resorbing agents it may act to release bone-resorbing cytokines from the osteoblast. To examine this hypothesis, we used serum-free conditioned media (CM) from SAOS II cells incubated with 10(-8) M h(1-74) PTH-RP for 48 h. Treated CM contained substantially more bone-resorbing activity (BRA) in the fetal-rat long-bone assay than CM from untreated cells (2.17 +/- 0.21 vs 1.38 +/- 0.16 fold stimulation over basal [f]; p less than 0.05]. After centrifugation and dialysis, 1 liter of treated CM contained a total BRA of 7102 ngeq b(1-34) PTH with a specific activity (SA) of 447 ngeq b(1-34) PTH/mg protein. Treated CM did not stimulate the ROS assay and the cytokines PGE2, TGF-alpha, EGF, GM-CSF and IL-1 were present in low concentrations. The BRA was heat sensitive. Ultrafiltration revealed that 97% of the BRA was in a 3-30 kD fraction. Further purification was achieved by sequential reverse phase HPLC and size exclusion-HPLC (SE-HPLC). A single fraction containing BRA from SE-HPLC was purified 277-fold to a SA of 123,810 ngeq b(1-34) PTH/mg protein and had an apparent MW of 9 kD. SDS-PAGE revealed 4 bands in this SE-HPLC fraction with 1 band at 9 kD unique to that fraction. PTH-RP may cause bone resorption in part by stimulating the release of a 9 kD protein from osteoblasts which is responsible for activating osteoclasts.

Biological Factors↗
PubMed1990

Persistence of sialodacryoadenitis virus in athymic rats.

To determine whether SDAV infection persists in athymic rats, weanling athymic rats and euthymic rats were inoculated intranasally with 10(4) TCID50 of SDAV and examined periodically for up to 90 days. Viral antigen and lesions characteristic of acute SDAV infection, including rhinotracheitis, bronchitis and sialodacryoadenitis, were detected in both groups of rats during the first week. In euthymic rats, tissues were under repair and viral antigen was undetectable by day 17, and tissues were histologically normal by day 31 except for mild focal dacryoadenitis. In athymic rats, viral antigen and chronic active inflammation of respiratory tract, salivary and lacrimal glands persisted through day 90. Inflammation and viral antigen also were observed in the transitional epithelium of the renal pelvis and urinary bladder as late as day 90. Virus was isolated from nasopharynx, lung, salivary gland and Harderian gland of athymic rats through day 90. All euthymic rats seroconverted to SDAV by day 6, whereas all athymic rats remained seronegative through day 31, and two of six were seropositive by day 90. As judged by seroconversion of contact sentinels, six of six athymic rats shed virus through 6 weeks, and five of six through 10 weeks. These results indicate that SDAV persists in athymic rats, and that normal T cell function is required for host defenses against SDAV.

Animals↗
PubMed1990

Infection of SDAV-immune rats with SDAV and rat coronavirus.

Infection of rats with sialodacryoadenitis virus (SDAV) or rat coronavirus (RCV) is acute and self-limiting, and elimination and control of either virus is based on the assumption that recovered rats are immune to reinfection. To test this hypothesis, we examined whether SDAV-immune rats could be infected with RCV or reinfected with SDAV. Sprague Dawley (SD) rats were inoculated intranasally with SDAV or with culture medium alone and serial SDAV antibody titers were obtained. Eleven months after inoculation, when antibody titers had stabilized, SDAV-immune and nonimmune rats were challenged with SDAV or RCV, and euthanatized 3 or 6 days later. SDAV-immune rats challenged with SDAV or RCV manifested acute rhinitis associated with virus antigen by 3 days after inoculation, but no lesions or antigen were subsequently found in the lower respiratory tract, salivary glands or lacrimal glands. There was also a marked anamnestic increase in antibody titer by 6 days after challenge. SDAV-immune rats challenged with SDAV or RCV also transmitted infection to nonimmune cage mates. This study indicates that 11 months after primary infection with SDAV, rats can be infected with SDAV or RCV, but that the severity of disease is significantly reduced.

Animals↗
PubMed1989

Transcriptional regulation of the parathyroid hormone-related peptide gene by glucocorticoids and vitamin D in a human C-cell line.

A parathyroid hormone-related peptide (PTHRP) has been identified in human tumors associated with the syndrome of humoral hypercalcemia of malignancy. The PTHRP and parathyroid hormone (PTH) genes appear to have arisen by duplication and to represent members of a gene family. PTHRP mRNAs have been demonstrated in a number of normal tissues, but little is known concerning the regulation of PTHRP gene expression in any site. We studied PTHRP gene expression in TT cells, a human C-cell line which also produces calcitonin and calcitonin gene-related peptide. We found that both the synthetic glucocorticoid, dexamethasone, and the active vitamin D metabolite, 1,25-dihydroxyvitamin D3, decreased steady-state PTHRP mRNA levels in TT cells in a time- and dose-dependent fashion. The dexamethasone effect was completely blocked by the glucocorticoid antagonist RU-486. 24,25-dihydroxyvitamin D3 was found to be inactive. Neither dexamethasone nor 1,25-dihydroxyvitamin D3 appeared to influence PTHRP mRNA stability in TT cells, and both agents were shown by nuclear transcription run-off assay to decrease PTHRP gene transcription. These findings indicate that the PTHRP gene is under the transcriptional control of glucocorticoids and vitamin D in a cell line with prototypical neuroendocrine features.

Calcitonin↗
PubMed1989

Clonal rat parathyroid cell line expresses a parathyroid hormone-related peptide but not parathyroid hormone itself.

A novel parathyroid hormone-related peptide has been identified in tumors associated with the syndrome of humoral hypercalcemia of malignancy. Subsequently, mRNAs encoding this peptide have been found to be expressed in a number of normal tissues, including the parathyroids. Using Northern blotting, RNase protection, and immunochemical techniques, we examined a clonal rat parathyroid cell line originally developed as a model system for studying parathyroid cell physiology. We found that this line expresses the parathyroid hormone-related peptide but not parathyroid hormone itself. Secretion of the parathyroid hormone-related peptide varied inversely with extracellular calcium concentration, but neither calcium nor 1,25-dihydroxyvitamin D3 appeared to influence steady-state parathyroid hormone-related peptide mRNA levels. This clonal line may prove to be an interesting system for studying the factors responsible for tissue-specific parathyroid hormone and parathyroid hormone-related peptide gene expression.

Animals↗
PubMed1989

Osteoblast-like cells secrete granulocyte-macrophage colony-stimulating factor in response to parathyroid hormone and lipopolysaccharide.

The cellular mechanism by which PTH and other osteotropic substances stimulate bone resorption is unclear. One hypothesis is that PTH-stimulated osteoblasts release cytokines which activate osteoclasts or osteoclast precursors. To examine whether cytokines are released by osteoblast-like cells in vitro, medium conditioned by a clonal rat osteosarcoma cell line 17/2.8 (ROS) was examined for mitogenic activity using a helper T lymphocyte line HT-2. This line proliferates in response to interleukin-2 (IL-2), IL-4, and granulocyte-macrophage colony-stimulating factor (GM CSF). Conditioned medium (CM) from untreated ROS cells caused proliferation of HT-2 cells. Treatment of ROS cells with PTH or lipopolysaccharide (LPS) caused a dose-dependent increase in the secretion of this mitogenic activity. To further define the nature of this mitogenic activity, we examined the effect of incubation of CM with neutralizing antibodies to IL-2, IL-4, and GM CSF. Mitogenic activity induced by both PTH- and LPS-treated ROS cell CM was completely inhibited by anti-GM CSF antibody, whereas there was no reduction in activity in the presence of antibodies to IL-2 or IL-4. Partial purification of both PTH- and LPS-treated CM using reverse phase HPLC resulted in a single peak of HT-2 mitogenic activity, which in both cases was completely inhibited by anti-GM CSF antibody. These findings suggest that PTH- and LPS-treated ROS cells secrete a T cell mitogenic activity which, by functional, serological, and biochemical criteria, is indistinguishable from GM CSF.

Animals↗
PubMed1988

Adenylate cyclase-stimulating, bone-resorbing and B TGF-like activities in canine apocrine cell adenocarcinoma of the anal sac.

Canine apocrine cell adenocarcinoma of the anal sac (APO-AS) is a spontaneously occurring tumor that causes humorally mediated hypercalcemia in 90% of cases. To further define the nature of the responsible mediator in APO-AS, we examined tumor extracts from five APO-AS and four control tumors for adenylate cyclase-stimulating activity (ACSA). All extracts from APO-AS contained potent ACSA, whereas the four control tumors did not. The ACSA extracted from one tumor demonstrated a dose response curve parallel to that of synthetic bovinePTH-(1-34) and was 80% inhibited by Nle8,18,Tyr34 bPTH-(3-34)amide at a concentration of 10(-5) M. Extracts from three APO-AS and three control tumors were also examined for in vitro bone-resorbing activity (BRA). All APO-AS contained significant BRA, stimulating resorption 1.47 to 2.13-fold over basal, whereas none of the control tumors stimulated resorption. Purification of one extract using C18 reverse-phase high pressure liquid chromatography (RP-HPLC) resulted in a single sharp peak of ACSA which was 400-fold purified compared with the initial extract. This pool also contained significant bone-resorbing activity, whereas none of the adjacent pools did. Purification of a second extract using sequential CN and C18 RP-HPLC followed by size exclusion HPLC resulted in material that was at least 10,000-fold purified, and showed co-purification of ACSA and B TGF-like activity.

Adenocarcinoma↗
PubMed1988

Identification of a cDNA encoding a parathyroid hormone-like peptide from a human tumor associated with humoral hypercalcemia of malignancy.

Humoral hypercalcemia of malignancy is a common paraneoplastic syndrome that appears to be mediated in many instances by a parathyroid hormone-like peptide. Poly(A)+ RNA from a human renal carcinoma associated with this syndrome was enriched by preparative electrophoresis and used to construct an enriched cDNA library in phage lambda gt10. The library was screened with a codon-preference oligonucleotide synthesized on the basis of a partial N-terminal amino acid sequence from a human tumor-derived peptide, and a 2.0-kilobase cDNA was identified. The cDNA encodes a 177 amino acid protein consisting of a 36 amino acid leader sequence and a 141 amino acid mature peptide. The first 13 amino acids of the deduced sequence of the mature peptide display strong homology to human PTH, with complete divergence thereafter. RNA blot-hybridization analysis revealed multiple transcripts in mRNA from tumors associated with the humoral syndrome and also in mRNA from normal human keratinocytes. Southern blot analysis of genomic DNA from humans and rodents revealed a simple pattern compatible with a single-copy gene. The gene has been mapped to chromosome 12.

Amino Acid Sequence↗
PubMed1988

In vitro adenylate cyclase-stimulating activity predicts the occurrence of humoral hypercalcemia of malignancy in nude mice.

A number of factors have been proposed as potential mediators of the syndrome of humoral hypercalcemia of malignancy (HHM), but to date no firm cause-and-effect relationship has been established. We attempted to establish such a relationship by determining whether the presence or absence of adenylate cyclase-stimulating activity (ACSA) in the media of cultured tumor cells predicted the occurrence of the syndrome of HHM when these cell lines were grown in nude mice in vivo. Conditioned media from 35 human renal carcinoma cell lines were surveyed for ACSA in the PTH-sensitive rat osteosarcoma 17/2.8 cell assay. 12 lines were positive (mean, 13.7-fold stimulation, range, 3.0 to 44.0), and 23 lines were negative (mean, 1.2-fold stimulation, range, 0.9 to 1.5). We were successful in establishing five of the positive and six of the negative lines in three to five nude mice per line. Mice implanted with the positive lines uniformly became hypercalcemic (mean serum calcium, 15.8 mg/dl), whereas mice implanted with the negative lines uniformly remained normocalcemic (mean serum calcium, 9.5 mg/dl), in spite of comparable mean tumor size. Acid-urea tumor extracts from each of four hypercalcemic animals contained potent in vitro ACSA (mean, 15.9-fold stimulation), while 5/5 extracts from normocalcemic animals did not (mean, 1.4-fold stimulation). Our study demonstrates that in this model system in vitro ACSA is a reliable predictive marker for HHM in vivo. Whether the protein responsible for this activity is also the mediator of the bone resorption seen in HHM remains to be demonstrated.

Adenylyl Cyclases↗
PubMed1988

Identification of transcripts encoding a parathyroid hormone-like peptide in messenger RNAs from a variety of human and animal tumors associated with humoral hypercalcemia of malignancy.

The syndrome of humoral hypercalcemia of malignancy (HHM) appears to be mediated in many instances by a parathyroid hormone-like peptide, which has recently been purified, sequenced, and cloned. Using a probe representing the coding region of the human PTH-like peptide, we examined by Northern analysis poly (A)+ RNA from a variety of human and animal tumors associated with HHM. Hybridizing transcripts were identified in mRNA from each of 12 human and each of four animal HHM-associated tumors, with a complex hybridization pattern observed in the human mRNAs and a relatively simple pattern observed in the animal mRNAs. Poly (A)+ RNA prepared from tumors of similar histological types unassociated with HHM failed to hybridize with the probe. Messenger RNA-dependent biological activity from the animal tumors was entirely eliminated in a hybridization-arrest experiment using a complementary oligonucleotide spanning the region of homology between human PTH and the PTH-like peptide. These findings indicate that the PTH-like peptide is associated with the syndrome of HHM in a wide spectrum of tumor types from a variety of mammalian species and that the PTH-like sequence in the proximal amino terminus of the peptide is highly conserved.

Amino Acid Sequence↗
PubMed1988

Humoral hypercalcemia of malignancy in canine lymphosarcoma.

Studies on the pathogenesis of hypercalcemia in canine lymphosarcoma have led to conflicting results. The biochemical and bone histomorphometric findings in canine lymphosarcoma were examined in 19 hypercalcemic and 17 nonhypercalcemic dogs with lymphosarcoma. Compared to the nonhypercalcemic group, the hypercalcemic dogs demonstrated an increase in fasting and 24-h calcium excretion, an increase in fractional phosphorus excretion, and a significant increase in nephrogenous AMP excretion. Plasma 1,25-dihydroxyvitamin D and immunoreactive PTH levels were equivalent in the two groups. Quantitative bone histomorphometry performed on iliac crest biopsies revealed increased parameters of bone resorption in those hypercalcemic dogs with no evidence of tumor at the biopsy site, without a compensatory increase in bone formation. Acid-urea tumor tissue extracts from eight hypercalcemic and six nonhypercalcemic dogs were examined for adenylate cyclase-stimulating activity (ACSA). All tumors from hypercalcemic dogs contained ACSA, whereas none of the tumors from nonhypercalcemic dogs had ACSA. Further purification of one tumor extract yielded an adenylate cyclase-stimulating protein which appeared to interact specifically with the PTH receptor. We conclude that in some cases, hypercalcemia in canine lymphosarcoma is mediated by a tumor-derived circulating bone-resorbing factor which is distinct from PTH. ACSA detected in tumor tissue appears to be a reliable marker for the syndrome in vivo. The role of this activity in the pathogenesis of the syndrome remains to be determined.

Adenylyl Cyclases↗