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Biomedical subjects

A T Cockett

Publications and source records attributed to A T Cockett.

At least 37 records · Page 2Linked to original sources
PubMed1996

Alteration of the hormonal bioactivity of parathyroid hormone-related protein (PTHrP) as a result of limited proteolysis by prostate-specific antigen.

OBJECTIVES: To discover whether the proteolytic activity of prostate-specific antigen (PSA) affects the structure and function of parathyroid hormone-related protein (PTHrP), as both are abundant components of human seminal plasma. METHODS: The ability of PTHrP to act as a substrate was studied by incubating a synthetic polypeptide, consisting of 34 amino acid residues of the amino-terminal domain of PTHrP, with purified PSA. The incubate was then analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, high-pressure liquid chromatography separation, amino-terminal peptide sequencing, and mass spectrometry. The physiologic effect of the proteolytic activity of PSA on PTHrP was studied by measuring any alteration in PTHrP (1-34)-induced elevation of cyclic adenosine monophosphate (cAMP) production by UMR-106 rat osteosarcoma cells in culture. All cell culture experiments were performed with PSA and PTHrP (1-34) at physiologic concentrations. RESULTS. Our data show that PSA proteolytically cleaves PTHrP (1-34) after either residue 22 or 23, generating three peptide fragments. Both cleavages occur carboxy terminally of a phenylalanine residue. The cAMP production in rat osteosarcoma cells, induced by the amino-terminal portion of PTHrP (1-34), as a result of its structural similarity with parathyroid hormone (PTH), was abated by PSA in a dose- and time-dependent fashion. In contrast, heat-inactivated PSA had no effect on cAMP production. CONCLUSIONS: Our study demonstrates that PTHrP is a substrate for PSA. The cleavage of the amino-terminal portion of PTHrP completely disrupts its ability to interact with the PTH/PTHrP receptor and thus inhibits its PTH-like activity. The proteolytic processing of PTHrP by PSA may play an important role in the post-translational/post-secretional regulation of prostatic PTHrP activities, which are believed to include regulation of prostate growth and differentiation.

Amino Acid Sequence↗
PubMed1996

Calcitonin receptor mRNA expression in the human prostate.

OBJECTIVES: A subpopulation of prostate neuroendocrine (NE) cells contain calcitonin (CT). It has been postulated that CT-producing cells in the prostate account for the high CT level in the semen, and may be involved in the regulation of other epithelial cells via a paracrine mechanism. The presence of CT binding sites in the plasma membrane fraction of prostate tissue has been demonstrated by radioligand binding assay. In the present study, we investigated the CT receptor gene expression in the human prostate, a key component of the autocrine/paracrine loop in the CT functional pathway. METHODS: Reverse transcription polymerase chain reaction (RT-PCR) was carried out to evaluate the CT receptor mRNA expression in normal prostate tissue. Subsequent DNA sequencing was used to verify RT-PCR amplified products and to determine the isoform of the receptor. To define the location of the CT receptor expression, nonradioactive in situ hybridization was performed with a digoxigenin-labeled probe complementary to the coding region of the CT receptor mRNA. A polyclonal antibody against CT was used to reveal the CT-secreting cells in the prostate. RESULTS: CT receptor MRNA expression was detected in the prostate tissue. Further analysis of the DNA sequence showed that CT receptor expressed in the prostate was the isoform without a 16-amino acid insert in the first intracellular domain. In situ hybridization revealed that CT receptor was present in the prostate NE cells. Immunocytochemical staining of mirror image sections showed that some CT-secreting cells also expressed CT receptor. CONCLUSIONS: CT receptor expression in the prostate, a key component in the CT functional pathway, is located in subsets of dispersed NE cells (CT secreting and CT nonsecreting), which indicates that prostate CT may play an important role in the autocrine/paracrine regulation of the prostate NE system.

Base Sequence↗
PubMed1995

Immunohistochemical localization of parathyroid hormone-related protein in prostatic intraepithelial neoplasia.

Parathyroid hormone-related protein (PTHrP) is a regulatory protein hormone that has been associated with normal fetal growth and differentiation as well as fetal calcium regulation. Parathyroid hormone-related protein has been implicated in a variety of carcinomas as a major factor in the development of humoral hypercalcemia of malignancy and may also play a role as an autocrine growth factor. In a previous immunohistochemical study we found that all prostatic adenocarcinomas (CAP) express PTHrP. In the current study, we evaluated PTHrP in prostate intraepithelial neoplasia (PIN) in radical prostatectomy specimens. A validated mouse monoclonal antibody, 9H7, raised against fragment 109-141 of the carboxy-terminus of PTHrP was used for immunostaining. The results generally showed negative to weak staining of normal and hyperplastic tissue and strong staining in PIN. The staining intensity was further evaluated by computer based image analysis. The relative optical density in PIN (9.24 +/- 9.05) was significantly (P = .008) higher than that in normal gland (.00 +/- 3.6). These findings suggest that PTHrP may be involved in the pathogenesis of prostatic dysplasia, and its immunohistochemical evaluation may have diagnostic use in the evaluation of PIN.

Adenocarcinoma↗
PubMed1995

Using repeated measures of symptom score, uroflowmetry and prostate specific antigen in the clinical management of prostate disease. Benign Prostatic Hyperplasia Treatment Outcomes Study Group.

Measurements of American Urological Association symptom score, peak urine flow rate and prostate specific antigen (PSA) are often followed over time in urological management. However, their interpretation is confounded by within-patient variability due to chance. Data from 2 clinical trials are used to examine the magnitude of this variation. When these measures are repeated at a short interval variation is modest and might easily be misinterpreted as a true change in patient condition. For example, approximately 20% of patients might be expected to have a chance increase or decrease in symptom score by at least 4.9 points, in peak urine flow rate by at least 4.1 ml. per second or in PSA by at least 1.6 ng./ml. Clinicians can use these data to help interpret repeated measures of these variables in patients, and can consider obtaining paired measurements to decrease the effect of chance variation when they plan on following them over time.

Aged↗
PubMed1994

Bladder irrigation specimens assayed by fluorescence in situ hybridization to interphase nuclei.

Bladder irrigation specimens provide a sampling of the entire bladder urothelium and are the most practical sample for longitudinally monitoring patients. This study presents cross-sectional fluorescence in situ hybridization (FISH) analyses with correlated DNA cytometry data on 76 patients monitored for recurrent bladder tumors. FISH probes complementary to centromeric satellite sequences for chromosomes 1, 7, 9, 11, 15, and 17 were used. Aberrations in copy number were observed for chromosomes 1, 7, 11, and 17 principally in patients with aneuploid tumors. Monosomy of chromosome 9 was observed in 39% of the diploid and 31% of the specimens with high hyperdiploid fraction. Significantly, 24% of patients with a history of bladder cancer but with no clinical evidence of disease exhibited monosomy of chromosome 9. This suggests a persistent and significantly large population of abnormal cells in the absence of clinical evidence of disease. Loss of chromosome 9 relative to DNA ploidy was observed in 24% of patients with no evidence of disease, in 59% of patients with tumor, and in 79% of patients with histologically confirmed transitional cell carcinoma, grades 1-3. Loss of chromosome 15 was also observed in a large percentage of patients. Loss of chromosome 15 was observed in 41% of specimens from patients in whom no tumor was seen, in 38% of specimens from patients with tumor, and in 67% of specimens from patients with histologically confirmed transitional cell carcinoma. Results of this study document the use of bladder irrigation specimens as a specimen source for FISH analyses.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗
PubMed1994

Overexpression of human epidermal growth factor receptor and c-erbB-2 by neuroendocrine cells in normal prostatic tissue.

OBJECTIVE: It has been suggested that prostatic neuroendocrine (NE) cells play an important role in the growth and differentiation of the prostate by secreting various neuropeptides and serotonin. However, the mechanism by which NE cells themselves are regulated is virtually unknown. In the present study we evaluated the expression of the human epidermal growth factor receptor (EGFR) family (HER) in prostatic NE cells. METHODS: Formalin-fixed, paraffin-embedded tissue sections from twenty radical prostatectomy specimens were immunostained with validated rabbit polyclonal antibodies raised against human EGFR and c-erbB-2, using the streptavidin-peroxidase enzyme conjugate method. RESULTS: A strong immunoreactivity was observed with both antibodies in the cytosol of a few epithelial cells. These cells frequently had a dendritic appearance and were located in the acini and ducts. The EGFR-positive cells were predominant in most cases. Double immunostaining revealed the colocalization of both antigens with chromogranin A, a polypeptide that is expressed by most NE cells. Moreover, EGFR and c-erbB-2 appeared to be colocalized as well as independently expressed by different subpopulations of NE cells. CONCLUSIONS: The results suggest that prostatic NE cells might be regulated by the HER protein family, probably, in a ligand-specific fashion. This is the first report identifying a potential pathway regulating prostatic NE cells.

Amino Acid Sequence↗
PubMed1994

Parathyroid hormone-related protein is expressed by prostatic neuroendocrine cells.

OBJECTIVE: Parathyroid hormone-related protein (PTHrP) is a regulatory peptide that has been associated with normal fetal growth and differentiation as well as the regulation of fetal calcium. In a variety of cancers, PTHrP has been implicated in the humoral hypercalcemia of malignancy. Recently, we demonstrated that all prostatic adenocarcinomas express PTHrP. In the present study, the localization of PTHrP and its mRNA in nonmalignant prostate tissue was assessed. METHODS: Formalin-fixed, paraffin-embedded prostatic tissues from 23 patients were evaluated. Immunocytochemistry (ICC) was performed by the streptavidin-peroxidase enzyme conjugate method using a monoclonal antibody, 8B12, generated against fragment 1-34 of the amino-terminal end of PTHrP. Nonradioactive in situ hybridization was carried out using a digoxigenin labeled single-stranded cDNA probe complementary to the sequence coding for PTHrP(15-120). RESULTS: PTHrP immunoreactivity was observed in the cytosol of a few epithelial cells. Double labeling and serial section ICC with 8B12 and a monoclonal antibody to chromogranin A (a generic neuroendocrine [NE] marker) revealed that PTHrP was present in a subpopulation of prostatic NE cells. In situ hybridization of mirror image sections demonstrated positive signals in prostatic NE cells in complete accordance with the ICC findings. CONCLUSIONS: The localization and production of PTHrP in prostatic NE cells suggest that this polypeptide may act in an endocrine-paracrine fashion involved in the prostatic growth and differentiation as well as the regulation of calcium in semen and seminal fluid.

Gene Expression↗
PubMed1994

Parathyroid hormone-related protein: a potential autocrine growth regulator in human prostate cancer cell lines.

OBJECTIVE: We recently demonstrated that parathyroid hormone-related protein (PTHrP) is widely expressed by human prostate cancer tissue, suggesting that PTHrP might be involved in the growth and development of prostate cancer. To study this further, the production of PTHrP and its biologic effect were investigated using human prostate cancer cell lines. METHODS: The cell lines used were one androgen-dependent cell line, LNCaP, and two androgen-independent cell lines, PC-3 and DU-145. PTHrP secreted by cancer cells was measured by radioimmunoassay. The effect of PTHrP on DNA synthesis in these cells was determined by thymidine incorporation assay. RESULTS: All cell lines secreted immunodetectable levels of PTHrP in the culture-conditioned media. PC-3 cells secreted significantly higher amounts than the other two cell lines. A synthetic peptide, PTHrP(1-34), stimulated thymidine uptake in PC-3 and DU-145 cells more than threefold the control under serum-free and steroid-free conditions, whereas LNCaP was not affected. However, in the presence of dihydrotestosterone, DNA synthesis of LNCaP cells was stimulated by PTHrP in a dose-dependent manner. Additionally, this PTHrP-induced DNA synthesis was completely neutralized by a validated mouse monoclonal antibody (8B12) raised against PTHrP(1-34). CONCLUSIONS: Our data suggest that PTHrP may play a significant role in the growth of prostate cancer by acting locally in an autocrine fashion.

DNA, Neoplasm↗
PubMed1994

The prostatic endocrine-paracrine (neuroendocrine) regulatory system and neuroendocrine differentiation in prostatic carcinoma: a review and future directions in basic research.

Endocrine-paracrine (neuroendocrine, amine precursor uptake and decarboxylation [APUD]) cells of the prostato-urethral region are serotonin and peptide containing regulatory cells, which are part of a dispersed neuroendocrine regulatory system also known as the APUD system. These cells most likely regulate growth and differentiation, as well as the secretory functions of the prostate. Prostatic carcinoma exhibits neuroendocrine differentiation in 3 forms: 1) small cell neuroendocrine carcinoma, 2) carcinoid-like tumors and 3) conventional prostatic adenocarcinoma with focal neuroendocrine differentiation. Small cell carcinoma and carcinoid-like tumors are rather rare (1 to 2% of all prostatic malignancies) and generally pursue an aggressive course. Focal neuroendocrine differentiation in adenocarcinoma is extensive in 10% of the cases and may be present in virtually all adenocarcinomas to a minor degree. There are conflicting studies on the prognostic significance of focal neuroendocrine differentiation in prostatic carcinoma, although several suggest a poor prognosis. The finding that serum neuroendocrine markers predict initial insensitivity to or the development of resistance to hormonal suppression therapy, coupled with the recent observation that androgen receptor is not expressed in neoplastic neuroendocrine cells suggests that neuroendocrine differentiation directly results in resistance to hormonal manipulation therapy. Neuroendocrine differentiation in prostatic carcinoma raises the possibility of innovative modes of treatment. Future directions of research should concentrate on the quantitative analysis of serotonin and various peptides in prostatic malignancy, since high levels of constitutive secretion may not be appreciated by immunocytochemistry, as well as analysis of tumors for receptors to neuroendocrine products, which are necessary for these products to have a functional role. Finally, specific subtypes of neoplastic cells with neuroendocrine differentiation based on serotonin and peptide profiles should be analyzed.

APUD Cells↗
PubMed1994

Androgen receptor immunostaining and its tissue distribution in formalin-fixed, paraffin-embedded sections after microwave treatment.

We used microwave (MW) oven heat treatment to unmask human androgen receptor (AR) immunostaining in formalin-fixed, paraffin-embedded tissue. Prostate tissue was used as an AR-positive control. Tissue sections were boiled in citrate buffer in a conventional MW oven for 30 min, followed by immunostaining with a validated murine monoclonal antibody (MAb), F39.4.1, raised against a peptide included in the N-terminal domain of the 100 KD human AR. AR immunostaining was localized to the nuclei of prostate secretory epithelial cells but was weak or absent in basal cells and of variable intensity in the stromal cells. Slides exposed to less than 10 min of MW heat treatment or none at all manifested no AR immunoreactivity. Tissue morphology was well preserved. Immunohistochemical determination of AR status in a wide variety of human tissues was consistent with that previously reported by others using frozen sections. MW heat treatment of tissue samples in an excellent method of localizing AR antigenicity, enabling immunohistochemical evaluation of AR status in formalin-fixed, paraffin-embedded material.

Adrenal Glands↗
PubMed1993

The androgen receptor status of neuroendocrine cells in human benign and malignant prostatic tissue.

Neuroendocrine (NE) cells containing neurosecretory granules, rich in various peptide hormones and biogenic amines, are components of the human prostate epithelium and prostatic adenocarcinomas. Neuroendocrine differentiation in prostatic adenocarcinomas has been associated with a poor prognosis and, following androgen withdrawal therapy, tumor cell populations have been observed to become enriched with NE cells. We assessed androgen receptor (AR) expression in NE cells in benign and malignant prostatic tissue using double-labeling immunocytochemistry with validated monoclonal antibodies to the AR and to chromogranin A (a generic NE marker). Neuroendocrine cells in benign and malignant prostatic tissue generally showed nuclear staining with AR. Some distinct AR-negative nuclei were observed in normal NE cells. In prostatic adenocarcinomas with extensive NE differentiation, a subpopulation of AR-negative NE cells was demonstrated. In conclusion, benign and malignant prostatic tissue contain both AR-positive and AR-negative NE cells that may have significance in regards to androgen-independent tumor growth and tumor progression.

Aged↗
PubMed1993

Immunohistochemical localization of parathyroid hormone-related protein in human prostate cancer.

Parathyroid hormone-related protein (PTHrP) is produced by a variety of malignant tumors and has been implicated as a major cause of humoral hypercalcemia of malignancy. Expression of PTHrP in prostate cancer tissue was studied immunohistochemically using 33 radical prostatectomy specimens from patients with clinically localized carcinoma of the prostate. None of these patients demonstrated hypercalcemia prior to the surgery. Acetone-methyl benzoate-xylene-processed, paraffin-embedded tissues were stained with a validated mouse monoclonal antibody to an amino acid fragment, PTHrP(109-141), using the streptavidin-peroxidase enzyme conjugate method. All cases (33 of 33; 100%) studied demonstrated some degree of immunoreactivity throughout the cytoplasm of the tumor cells, but immunostaining was absent from inflammatory and stromal cells. The intensity of the staining appeared to directly correlate with increasing tumor grade. The widespread immunohistochemical localization of PTHrP in carcinoma of the prostate suggests that PTHrP may play some local role in the growth of transformed cells in the prostate. Furthermore, overexpression of PTHrP may be a possible marker to evaluate the malignant potential of carcinoma of the prostate.

Aged↗
PubMed1993

Insulin-like growth factor I: action and receptor characterization in human prostate cancer cell lines.

The role of insulin-like growth factor I (IGF-I) in the growth and development of prostate cancer was studied using established human prostate cancer cell lines. Under steroid and growth factor-free culture conditions, IGF-I significantly stimulated the androgen-independent cell lines PC-3 and DU-145 to incorporate [3H]thymidine into DNA, while the androgen-dependent cell line, LNCaP, was not affected. However, in the presence of dihydrotestosterone (DHT), DNA synthesis of LNCaP cells was stimulated by IGF-I in a dose-dependent manner. None of the cell lines tested secreted an immunoreactive level of IGF-I into their conditioned medium. Characterization of receptors by ligand binding assays revealed that all prostate cancer cell lines tested express specific binding sites for IGF-I with similar dissociation constants (0.23-0.39 nM). Crosslinking studies supported the suggestion that 125I-IGF-I was bound to a receptor on these cells. The IGF-I receptor concentrations of androgen-independent cell lines were significantly higher than those of the androgen-dependent cell line. Androgen appeared to affect neither the expression of IGF-I receptors nor the secretion of IGF-I. The results suggest that IGF-I may play an important role in stimulating the growth and progression of prostate cancer.

Binding, Competitive↗
PubMed1993

Characterization of insulin-like growth factor I binding sites in human bladder cancer cell lines.

The role of insulin-like growth factor I (IGF-I) in the growth and development of bladder cancer cells was investigated using cultured human cell lines representing differentiated (RT-4, 5637) or undifferentiated (T-24, J-82, TCC-SUP) transitional cell carcinoma (TCC). In the presence of 2% serum, IGF-I significantly stimulated the growth of all cell lines. The proliferation of T-24, 5637, and RT-4 cells was more sensitive to IGF-I than that of J-82 and TCC-SUP cells. [125I]IGF-I binding to 5637 and J-82 cells was significantly higher than that to T-24 and TCC-SUP cells (P < 0.001). RT-4 cells possessed the lowest binding capacity among the cell lines tested. Scatchard analysis of [125I]IGF-I binding to four of the five cell lines indicated a single binding site for IGF-I, with apparent dissociation constants (Kd) of 1.27, 1.18, 1.34, and 1.39 nmol/l for TCC-SUP, J-82, 5637, and T-24, respectively. Therefore, the difference observed in [125I]IGF-I binding among the bladder cancer cell lines was attributed to the difference of IGF-I binding sites and not to a change in receptor binding affinity. Cross-linking studies supported the suggestion that [125I]IGF-I was bound to a receptor on these cells. The results indicate that cultured human bladder cancer cells contain functional IGF-I receptors. A differentiated cell line, RT-4, possesses significantly fewer IGF-I receptors than other cell lines. This suggests that the overexpression of IGF-I receptor may reflect the malignant potential of bladder cancer cells.

Binding Sites↗
PubMed1993

Relationship of neuroendocrine cells of prostate and serotonin to benign prostatic hyperplasia.

Neuroendocrine (NE) cells containing neurosecretory granules, rich in various peptide hormones and biogenic amines such as serotonin (5-HT), are components of the human prostate epithelium. The NE cells probably subserve a paracrine or local regulatory role in both prostatic growth and differentiation as well as the exocrine secretory process. Neuroendocrine cells may be involved in the etiology of benign prostatic hyperplasia (BPH). In this study the number of NE cells in areas of BPH was compared with normal tissue using 5-HT immunocytochemistry. In addition, using high-performance liquid chromatography with electrochemical detection (HPLC-ECD), tissue levels of 5-HT and its metabolite 5-hydroxy-indoleacetic acid (5-HIAA) were analyzed in prostatic tissue extracts including 25 cases of BPH and 16 cases of normal tissue verified by adjacent histologic sections. Compared with normal prostate our results demonstrated a marked decrease in 5-HT immunoreactive NE cells in the vast majority of larger hyperplastic nodules of BPH. These findings were corroborated by quantitative analysis where a significant reduction in the tissue 5-HT levels in BPH (0.539 +/- 0.09 SE) compared with normal (1.75 +/- 0.22 SE) (p < 0.05) was found. When smaller nodules of BPH were studied, abundant NE cells, equal or increased in number compared with those in adjacent normal prostatic tissue, were seen. The small apparently developing BPH nodules and ductal-like structures contained NE cells which may be growth foci near the periphery of some hyperplastic nodules. These findings particularly in small hyperplastic nodules suggest that NE cells and their products involved in controlling cell proliferation through a paracrine hormonal mechanism and may be involved in the pathogenesis of BPH. Serotonin (5-HT) and NE peptides may represent that elusive local "missing link" often alluded to in various theories relating to the development of early nodular hyperplasia in BPH.

Aged↗
PubMed1993

Role of extracorporeal shock-wave lithotripsy in treatment of urolithiasis in patients with medullary sponge kidney.

In an effort to identify the role of extracorporeal shock-wave lithotripsy (ESWL) in the treatment of patients with medullary sponge kidney, a retrospective review of our experience with the Dornier HM-3 lithotriptor is presented. From 1986-1991 17 patients with medullary sponge kidney ranging in age from twenty-eight to fifty-eight years received 31 treatments to a total of 24 renal units. Follow-up ranged from four to fifty-six months with an average follow-up of twenty-five months. ESWL appears to have a role in the treatment of symptomatic urolithiasis in patients with medullary sponge kidney. There may also be some benefit in treating parenchymal calcifications. No significant side effects from the treatment were noted.

Adult↗