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PubMed1993

Kainic acid seizures in the rat: differential expression of chromogranin A, carboxypeptidase H and peptidylglycine alpha-amidating monooxigenase in subfields of the hippocampal formation.

Using in situ hybridization histochemistry concentrations of mRNAs encoding chromogranin A (ChA), carboxypeptidase H (CPH) and peptidylglycine alpha-amidating monooxigenase (PAM) have been investigated in the hippocampus after kainic acid (KA)-induced limbic seizures in the rat. Increased concentrations (by 150%) of ChA and CPH mRNAs were found in the granule cell layer 24 h after KA injection. At the same time PAM mRNA levels were only slightly elevated (by 50%). Whereas the increases in CPH and PAM transcripts were only transient, ChA mRNA concentrations in the granule cell layer were elevated up to 2 months after the initial seizures. In contrast, in the pyramidal cell layers of all hippocampal subfields (CA1 to CA3) ChA mRNA concentrations were significantly reduced (by 40-70%) 1-60 days after KA. PAM and CPH messages were slightly reduced in the pyramidal cell layer of CA1 but not in CA2 and CA3. The experiments demonstrate that KA-induced limbic seizures cause sustained changes in the expression of ChA mRNA. At the same time the expression of two enzymes involved in post-translational processing of neuropeptides, PAM and CPH, becomes only transiently altered. Synthesis of ChA may be regulated differently in the strata granulosum and pyramidale during epileptic seizures.

Animals↗

PubMed1990

Effects of cytotoxic prostaglandin, delta 12-PGJ2 on protein synthesis and cytoskeleton in transformed epidermal cells in culture.

Cyclopentenone prostaglandins (PGs) such as delta 12-PGJ2 and PGA are potent inducers of growth inhibition in a variety of cultured cells, including epidermal cells. These PGs are actively transported into cells by a specific carrier on cell membrane and accumulate in cell nuclei with binding to nuclear protein. To clarify the mechanism of cytotoxicity of these PGs in epidermal cells, we examined the effects of delta 12-PGJ2 on protein synthesis and cytoskeleton in the PAM 212 transformed mouse epidermal cell line. Cycloheximide at 1 microgram/ml culture medium exhibited a protective effect on cell growth inhibition of PAM 212 cells by delta 12-PGJ2. The analysis of cell lysate protein patterns by SDS-polyacrylamide gel electrophoresis revealed that 12-h incubation with delta 12-PGJ2 increased the amount of 70 kD protein in PAM 212 cells. The amount of 70 kD protein in delta 12-PGJ2-treated cells was markedly decreased by cotreatment with cycloheximide. This 70 kD protein was also induced in PAM 212 cells with treatment at 43 degrees C for 90 min, indicating that this synthesized protein belongs to the heat shock protein. The addition of delta 12-PGJ2 to confluent PAM 212 cells resulted in the disappearance of action filament, as visualized by fluorescent labeled phallacidine, but in contrast, keratin filament appeared to be intact during 12-h incubation with delta 12-PGJ2 at a concentration of 5 micrograms/ml culture medium. These results suggest that the cytotoxicity of cyclopentenone PGs is at least in part due to induction of the synthesis of some protein(s), probably one of the heat shock proteins, and the damage to the actin filament in transformed cultured epidermal cells.

Actin Cytoskeleton↗

PubMed1996

Human middle latency auditory evoked magnetic fields.

The magnetic equivalents of SN10, Po, Na, Pa, Nb and Pb (SN10m, Pom, Nam, Pam, Nbm, and Pbm) in short and middle latency auditory evoked potentials were measured with a 7-channel DC superconducting quantum interference device (SQUID). The sources of Pom, Nam, Pam, Nbm and Pbm responses were estimated to be located in the auditory cortex, while the source of SN10m was considered to be in a deeper part of the brain. In addition, the source of Pam was estimated to be in the vicinity of the moving N100m source. The source of Pbm was considered to be in a separate area, anterior to the source of Pam and N100m, which suggested that source of Pam was located in the primary auditory cortex, while the source of Pbm was located in the secondary auditory cortex. The source of N100m was considered to spread from the primary auditory cortex to the secondary auditory cortex.

Adult↗

PubMed1995

Neurotransmitter suppression of the in vitro generation of a cytotoxic T lymphocyte response against the syngeneic MOPC-315 plasmacytoma.

We have previously shown that, as a consequence of low-dose melphalan (L-phenylalanine mustard (L-PAM) therapy, the hitherto immunosuppressed spleen cells from BALB/c mice bearing a large MOPC-315 tumor (in contrast to spleen cells from normal mice) acquire the ability to generate a greatly enhanced anti-MOPC-315 cytotoxic T lymphocyte (CTL) response upon in vitro stimulation with MOPC-315 tumor cells. Here we show that the catecholamines norepinephrine, epinephrine, and isoproterenol suppressed the in vitro generation of anti-MOPC-315 cytotoxicity by spleen cells from mice that had just completed the eradication of a large MOPC-315 tumor following low-dose L-PAM therapy (L-PAM TuB spleen cells), as well as by spleen cells from normal mice. In contrast to the marked suppression obtained with catecholamines, the cholinergic agonist carbachol had no effect on the in vitro generation of splenic anti-MOPC-315 cytotoxicity. The inhibitory effect of the catecholamines was "mimicked" by the membrane penetrating analog of cAMP, dibutyryl-cAMP, and by cholera toxin at concentrations that stimulate the endogenous production of cAMP. The beta-adrenergic receptor antagonist propranolol did not block norepinephrine-induced inhibition of the generation of anti-MOPC-315 cytotoxicity by either normal or L-PAM TuB spleen cells. Since the curative effectiveness of low-dose L-PAM therapy for MOPC-315 tumor bearers requires the participation of CD8+ T cells that exploit a CTL response in tumor eradication, it is conceivable that norepinephrine may reduce the therapeutic outcome of low-dose chemotherapy by inhibiting the acquisition of CTL activity.

Animals↗

PubMed1995

Breast cancer after breast augmentation with silicone implants.

BACKGROUND: Although epidemiological studies have failed to demonstrate an increased incidence of breast cancer in women who had undergone prior prosthetic augmentation mammoplasty (PAM), it has been reported that when breast cancer arises in this group it presents mostly in a palpable form and at a more advanced stage. This is thought to be secondary to suboptimal mammographic evaluation caused by the masking effect of the implant. This study was undertaken to determine, in our experience, whether breast cancer arising in women who had undergone PAM could be detected in a prepalpable form by mammography and whether it presented at a more advanced stage as compared with nonaugmented women with breast cancer. METHODS: The charts of 22 patients, treated by at least one of the authors, in whom 23 breast cancers developed after PAM (group A) were retrospectively reviewed. The comparison groups consisted of 611 nonaugmented patients who underwent 636 procedures for the treatment of primary breast cancer at our institution (group B) and the surveillance, epidemiology, and end results (SEER) data (group C). Parameters studied were mode of detection, tumor size, axillary lymph node involvement, and histopathology. RESULTS: No significant differences between the groups were found in mean tumor size (group A vs. group B), the incidence of preinvasive cancer (group A vs. group B) or axillary lymph node involvement (group A vs. group B and group A vs. group C). Breast-preserving surgery was performed significantly less in augmented patients (group A vs. group B). CONCLUSION: We conclude that prepalpable and preinvasive breast cancer can be detected in the PAM patient by mammography and that the stage of presentation in this group is not significantly different than in nonaugmented patients. Total mastectomy is preferred over breast-preserving procedures for the treatment of breast cancer in the PAM patient.

Adult↗

PubMed1993

Predicting survival from in-hospital CPR: meta-analysis and validation of a prediction model.

OBJECTIVE: To better clarify patient factors that predict survival from in-hospital cardiopulmonary resuscitation (CPR), using two methods: 1) meta-analysis and 2) validation of a prediction model, the pre-arrest morbidity (PAM) index. DESIGN: Meta-analysis of previously published studies by standard techniques. Retrospective chart review of validation sample. SETTING: University-affiliated teaching hospital. PATIENTS/PARTICIPANTS: Meta-analytic sample of 21 previous studies from 1965-1989. The validation sample consisted of all patients surviving resuscitation from the authors' hospital during the period September 1986 to January 1991. A matched sample of patients who did not survive from the same time period was used as the comparison group. INTERVENTIONS: None. MEASUREMENTS AND MAIN RESULTS: The strongest negative predictors of survival, by meta-analysis, were renal failure (r = 0.088, p < 0.0002), cancer (r = 0.08, p < 0.0002), and age more than 60 years (r = 0.063, p < 0.006). Sepsis (r = 0.046, p < 0.02), recent cerebrovascular accident (CVA) (r = 0.038, p < 0.04), and congestive heart failure (CHF) class III/IV (r = 0.036, p < 0.05) were weaker negative predictors. Presence of acute myocardial infarction (AMI) was a significant positive predictor of survival (r = 0.15, p < 0.0001). The PAM score was highly predictive of survival in a logistic regression model (p < 0.0003, R2 = 9.6%). No patient who survived to discharge had a PAM score higher than 8. CONCLUSION: Meta-analysis reveals that the most significant negative predictors of survival from CPR are renal failure, cancer, and age more than 60 years, while AMI is a significant positive predictor. The PAM index is a useful method of stratifying probability of survival from CPR, especially for those patients with high PAM scores, who have essentially no chance of survival.

Age Factors↗

PubMed1987

From growth factor dependence to growth factor responsiveness: the genesis of an alveolar macrophage cell line.

A rat pulmonary alveolar macrophage (PAM) cell line (NR8383) was initiated in culture in the presence of a gerbil lung cell conditioned medium (GLCM), and has been propagated continuously for over 36 mo. When examined at different times throughout this in vitro period, NR8383 exhibited characteristics typical of macrophages: (a) Zymosan ingestion was seen in 90 to 98% of the cells examined; (b) Pseudomonas aeruginosa phagocytosis in 50 to 80%; (c) Nonspecific esterase activity in greater than 95%. During the first 6 mo., the PAM replicated with doubling times approximating 15 to 20 d. Throughout this period, GLCM dependence was evident. After 27 wk in vitro, NR8383 replication increased markedly, and within 2 wk, the doubling time was less than 48 h. NR8383 was readily monitored by [3H]thymidine (TdR) blastogenesis assay. In the presence of GLCM uptake of [3H]TdR was fivefold greater than in control cultures. Adherence and growth kinetics were effectively controlled by modulation of GLCM or serum content in culture medium. It was demonstrated that PAM growth factor(s) is ubiquitous, not species-specific, and under certain conditions may be derived from "endogenous" sources of persisting non-PAM populations within the parent, uncloned line NR8383. Cloned progeny remain devoid of non-PAM "feeder" cells, but retain macrophage properties, including interleukin-1 secretion, Fc receptors, and H2O2 production.

Animals↗

PubMed2000

Comparative study on three chemotherapeutic regimens for the treatment of advanced epithelial ovarian cancer.

To investigate the best first-line chemotherapy regimen for the treatment of advanced epithelial ovarian cancer (AEOC), the efficacy of three chemotherapy regimens for treatment of the patients with AEOC in our hospital during Jan. 1992-Jan. 1999 was retrospectively analyzed. The therapeutic effects were compared with the supplement of Melphalan + Hexamethylme (PAM + HMM), cisplatin + adriamycin + cyclophosphamide or isofamide (PAC) or cisplatin + cyclophosphamide or isofamide (PC), Taxol + cisplatin (TP) combined chemotherapy after cytoreductive surgery. The results showed that the overall effective rate of TP was significantly higher than that of PAM + HMM (P < 0.05); The complete remission rate of TP was significantly higher than that of PAM + HMM and PAC or PC (all P < 0.05); The 2-year survival rate free of tumor of TP was obviously higher than that of PAM + HMM and PAC or PC (all P < 0.05). It was concluded that the therapeutic effect of TP regimen in the treatment of AEOC was better than PAM + HMM and PAC or PC and TP regimen could be recommended currently as the preferred first-line one for the treatment of AEOC.

Adult↗

PubMed2004

Open-nucleus breeding strategies compared with population-wide positive assortative mating: I. Equal distribution of testing efforts.

Positive assortative mating (PAM) can enhance the additive genetic variance in a breeding population(BP). This increases the potential for gains in the production population (PP, selected subset of the BP) for recurrent selection programs in forest trees. The assortment of mates can be either: (1) by individual tree rank across the whole BP (PAM), or (2) trees of similar rank can be merged into larger hierarchical groups and then mated randomly within group ("open"-nucleus breeding,NB). The objective of this study was to compare PAM and NB in quantitative terms. The NB simulation model assumed two tiers (nucleus, main) with unrestricted migration between the tiers. Clonal tests were used to predict breeding values and test resources per mate were kept constant for all mates. Both gain and diversity were combined into a single selection criterion, "group-merit selection." Alternatives were compared over five breeding cycles by considering genetic gain and diversity in a selected PP established in a seed orchard. The assortment of mates in both alternatives enhanced additive variance and increased the additive effect in the BP, leading to additional gain in the PP. Gains generated under PAM always exceeded gains under NB. Thus, the main message from this study is that PAM in both the short- and long-term results in more gain at any target level of diversity in the PP (the breeder's target) than is achieved by the NB alternative. The optimum size of the nucleus varies with the desired level of seed orchard diversity. At lower target diversity, smaller nucleus sizes are favorable, while larger sizes result in more gain when seed orchard diversity is considered more important.

Breeding↗

PubMed2002

Gene cloning, overexpression and biochemical characterization of the peptide amidase from Stenotrophomonas maltophilia.

The peptide amidase (Pam) from the gram-negative bacterium Stenotrophomonas maltophilia catalyzes predominantly the hydrolysis of the C-terminal amide bond in peptide amides. Its gene ( pam) was isolated by Southern hybridization using a DNA probe derived from the known N-terminal amino acid sequence. Pam is a member of the amidase signature family and was identified as a periplasmic protein by an N-terminal signal peptide found in the gene. The processed protein consists of 503 amino acids with a molecular mass of 53.5 kDa. The recombinant enzyme with a C-terminal His(6) tag has a monomeric structure and its isoelectric point is 6.3. The dipeptide amide L-Ala- L-Phe-NH(2) is hydrolyzed in the absence of cofactors to L-Ala- L-Phe-OH and ammonia with V(max)=194 U/mg and K(m) <0.5 mM. The natural function of Pam remains unclear. Chymostatin ( K(i)<0.3 microM) and Pefabloc SC ( K(i) not determined) were identified as inhibitors. When the gene was expressed in Escherichia coli on a 12-l scale, the specific activity in the crude extract was 60 U/mg, compared to 0.24 U/mg in S. maltophilia. In the expression system, Pam made up about 31% of the total soluble cell protein. From 75 g wet cells, 2.1 g of >95% pure enzyme was obtained, which corresponds to a total activity of 416,000 units.

Amidohydrolases↗

PubMed2006

Effect of antibiotic heterogeneity on the development of infections with antibiotic-resistant gram-negative organisms in a non-intensive care unit surgical ward.

BACKGROUND: Heterogeneous antibiotic use has been suggested to limit the emergence of resistance, but determining the optimal strategy is difficult. METHODS: We developed a new strategy, termed "periodic antibiotic monitoring and supervision" (PAMS) program in a non-ICU surgical ward. The 2-year prospective study was divided into a 1-year observation period and a 1-year PAMS period. The use of four major classes of antibiotics in empirical therapy for Gram-negative rod (GNR) infections was supervised. During the PAMS program, recommended, restricted, and off-supervised classes of antibiotics were changed every 3 months according to the usage pattern of the antibiotics in the preceding term. RESULTS: Cefepime (45.5%) and imipenem/cilastatin (39.4%) were the most common antibiotics of choice during the observation period. The use of these antibiotics decreased significantly during the PAMS period, and that of fluoroquinolones and extended-spectrum penicillin/beta-lactamase inhibitor increased (4.8% vs. 21.4% and 2.4% vs. 21.4%, P<0.01 respectively). Outcome analysis demonstrated a tendency toward reduction in the incidence of resistant GNR infections (P=0.079) and that of Pseudomonas aeruginosa (P=0.053). The incidence of resistant Gram-positive core infections did not decrease. Analysis of antibiotic susceptibility to GNR revealed no significant beneficial results for any antibiotics. CONCLUSIONS: As significant changes were not observed, the PAMS program is not generally applicable and heterogeneous antibiotic use as a way of reducing infections with resistant GNR in non-ICU surgical wards was not established.

Anti-Bacterial Agents↗

PubMed1999

A pilot study of melphalan, tumor necrosis factor-alpha and 41.8 degrees C whole-body hyperthermia.

PURPOSE: To evaluate the feasibilitv of sequencing (based on preclinical modeling) tumor necrosis factor-a (TNF) at two dose levels with melphalan (L-PAM) and 41.8 C whole-body hyperthermia (WBH) for 60 min. PATIENTS AND METHODS: Nine patients with refractory cancer were treated from October 1995 to June 1997. The study encompassed a total of 20 trimodality treatment courses. Three patients were treated at TNF dose level I (50 microg/m2) and six patients were treated at TNF dose level II (100 microg/m2). TNF was delivered as a 24-h intravenous infusion, 48 h prior to the combination of L-PAM and WBH; L-PAM was given over 10 min at target temperature at a dose of 17.5 mg/ m2 based on a previous phase I WBH/L-PAM trial. WBH was administered with an Aquatherm radiant heat device. RESULTS: Myelosuppression was the major toxicity associated with therapy, but there were no instances of bleeding or neutropenic fevers. Grade 3 thrombocytopenia was seen with 15% of treatments. Regarding absolute neutrophil count, 15% of treatments were associated with grade 3 toxicity, and 45% with grade 4 toxicity, and regarding white blood cell count, 50% of treatments were associated with grade 3 toxicity and 10% with grade 4 toxicity. The myelosuppression observed was equivalent to that seen in our earlier phase I study of WBH and L-PAM (without TNF). Only mild toxicities (grade 1 or 2) were associated with TNF; these were seen with <25% of treatments and included nausea, vomiting, diarrhea, fevers, and headache. There were no instances of hypotension. There was no relationship between toxicities observed and the two TNF dose levels. Mild WBH toxicities were seen with less than 15% of treatments; these included nausea, vomiting, and herpes simplex I. Responses included two complete remissions (malignant melanoma, TNF dose level I; breast cancer, TNF dose level II), and two disease stabilizations (both malignant melanoma, TNF dose level I). CONCLUSION: We conclude that the combination of TNF, L-PAM, and WBH is well tolerated at the dose levels studied. The clinical results justify further clinical investigation for this trimodality treatment approach.

Adult↗

PubMed2002

Increase in serum IgE levels following injection of syngeneic keratinocyte extracts in BALB/c mice.

Increased serum IgE levels are found in atopic diseases, especially atopic dermatitis. It is generally accepted that the high serum IgE levels are increased by exposure to exogenous antigens. However, exposure to antigens in the absence of proper adjuvants does not usually increase serum IgE. Here, we studied the involvement of skin factor(s) in increasing serum IgE. The soluble fractions were obtained from two keratinocyte cell lines, PAM 212 and KCMH-1, derived from BALB/c and CBA/J mice, respectively. These were injected subcutaneously into parental mice strains at 2-week intervals. The levels of serum immunoglobulins and the release of IgE in vitro from spleen cells were measured by ELISA. The biological activity of the increased serum IgE was assessed by sensitization of RBL-2H3 mast cells with serum. Six injections of PAM 212-derived fraction (PAM extract) consistently increased the serum levels of IgE, IgG1 and IgG2b, but not those of IgM, IgG2a, IgG3 or IgA in BALB/c mice. The high serum levels of IgE were sustained for at least 10 weeks after one or two injections of PAM extract. When RBL-2H3 cells were sensitized with the serum from mice injected with PAM extract, they released histamine in response to anti-IgE. Moreover, spleen cells of these mice produced significantly higher amounts of IgE than those of uninjected mice in the presence of IL-4 and anti-CD40 antibody. On the other hand, no significant increase in immunoglobulins was observed following injection of the soluble fraction from KCMH-1 cells in CBA/J mice. These results suggest that the release of the soluble fraction of keratinocytes and its exposure to immunologically competent cells may induce an increase in serum IgE levels by a Th2-dominant mechanism.

Adjuvants, Immunologic↗

PubMed2006

The extract of syngeneic keratinocytes enhances IgE production from BALB/c mouse splenic lymphocytes in vitro.

The increase of serum IgE levels is closely associated with atopic dermatitis. We have previously revealed that cellular extract of PAM212 cells (PAM-extract), BALB/c mouse keratinocyte cell line, induced a remarkable increase of serum IgE levels, in vivo, when subcutaneously injected into BALB/c mice. However, precise mechanism of IgE-increasing activity was unclear. To elucidate the mechanism of IgE-increase in sera of BALB/c mice induced by PAM-extract, we explored the direct influence of PAM-extract on immunoglobulin production and class-switching in the culture of splenic lymphocytes and purified B-cells, in vitro. Splenic lymphocytes or purified B-cells obtained from BALB/c mice were cultured with various combinations of IL-4, anti-CD40 antibody, and PAM-extract for 7 days. IgE and IgG concentrations of culture supernatants were measured by ELISA. Epsilon germ-line transcriptions were assessed by RT-PCR from the cultured cells. IgE and IgG concentrations in culture supernatant of splenic lymphocytes were increased by an addition of PAM-extract in the presence of both IL-4 and anti-CD40 antibody. Epsilon germ-line transcript was also induced in parallel to the increase of IgE production. Similar results were obtained when purified B-cells were employed in stead of whole splenic lymphocytes. The enhancement of IgE production in vitro was also observed, when splenic lymphocytes of CBAj mouse were cultured with cellular extract of KCMH-1 cells, CBAj mouse keratinocyte cell line. The cellular extract of keratinocyte promotes immunoglobulin class-switching to IgE and IgE production from mouse splenic B-cells in an IL-4- and CD40-stimuli-dependent manner. Such enhancement may account for the increase of serum IgE in patients with dermatitis in association with a Th2 microenvironment.

Animals↗

PubMed2004

Cost analysis of secondary prophylaxis with oral clodronate versus pamidronate in metastatic breast cancer patients.

OBJECTIVE: We compared the direct medical costs of secondary prophylaxis with bisphosphonates (BPs) in bone metastases (BMs) of breast cancer (BCa) from a payer perspective. PATIENTS AND METHODS: This study adopted an incidence-based chart review of consecutive BCa patients with BMs who received prophylactic treatment with orally administered (po) clodronate (CLODpo group), or intravenously administered (iv) pamidronate (PAM group) in 1997 at two large oncology centers in Toronto, Ontario. We evaluated the difference in costs of management of patients among the CLODpo and PAM groups using an intent-to-treat analysis from diagnosis of BMs to death, or last follow-up. The results are presented as observed mean and average lifetime (including terminal care) costs per patient. RESULTS: The observed mean costs in the PAM and CLODpo groups were 49,472 dollars and 50,307 dollars (2002 Canadian dollars), respectively. The difference in costs between the CLODpo (n=34) and PAM (n=18) groups was not significant (P=0.64), and remained robust after sensitivity analyses. The corresponding average lifetime costs were 65,677 dollars in the CLODpo group and 61,254 dollars in the PAM group. Inpatient and terminal care were the major cost drivers, comprising 45% and 25% of overall costs. Of all hospitalizations, 46% were associated with complications from BMs. CONCLUSIONS: Our analysis, which was based on a convenience sample, failed to reveal a statistically significant difference in the observed mean costs between groups of patients who initiated treatment with po clodronate versus iv pamidronate. The cost estimates from this study can be used for future corroborative economic analyses.

Antimetabolites↗

PubMed2005

(Z,Z)-6,9-Heneicosadien-11-one: major sex pheromone component of painted apple moth, Teia anartoides.

(Z,Z)-6,9-Heneicosadien-11-one (Z6Z9-11-one-21Hy) was identified as the major sex pheromone component of the painted apple moth (PAM), Teia anartoides (Lepidoptera: Lymantriidae), on the basis of (1) comparative gas chromatographic-electroantennographic detection (GC-EAD) analyses, GC-mass spectrometry (MS), high-performance liquid chromatography (HPLC)-MS, and HPLC-UV/visible spectroscopy of pheromone gland extracts and authentic standards; (2) GC-EAD analyses of effluvia of calling females; and (3) wind tunnel and field trapping experiments with a synthetic standard. In field experiments in Australia, synthetic Z6Z9-11-one-21Hy as a single component attracted male moths. Wind tunnel experiments suggested that a 4-component blend consisting of Z6Z9-11-one-21Hy, (6Z,9R,10S)-cis-9,10-epoxy-heneicosene (Z6-9R10S-epo-21 Hy), (E,E)-7,9-heneicosadien-6, 11-dione (E7E9-6,11-dione-21Hy), and 6-hydroxy-(E,E)-7,9-heneicosadien-11-one (E7E9-6-ol-11-one-21Hy) (all present in pheromone gland extracts) might induce more males to orient toward, approach, and contact the source than did Z6Z9-11-one-21Hy as a single component. Additional experiments are needed to determine conclusively whether or not Z6-9R10S-epo-21Hy, E7E9-6,11-dione-21Hy, and E7E9-6-ol-11-one-21Hy might be minor sex pheromone components of PAM. Moreover, attractiveness of synthetic pheromone and virgin PAM females needs to be compared to determine whether synthetic pheromone could replace PAM females as trap baits in the program to monitor eradication of exotic PAM in New Zealand.

Animals↗

PubMed1985

Further characterization of the peptidyl alpha-amidating enzyme in rat anterior pituitary secretory granules.

In previous studies we have demonstrated a secretory granule-associated peptide alpha-amidation activity in rat anterior, intermediate, and posterior pituitary. This activity is capable of converting 125I-labeled synthetic D-Tyr-Val-Gly to labeled D-Tyr-Val-NH2, and requires ascorbic acid, CuSO4, and molecular oxygen for optimal activity. Because of the requirement for peptides with COOH-terminal glycine residues, and cofactor requirements similar to monooxygenases such as dopamine beta-monooxygenase, we have proposed that the alpha-amidating enzyme be named peptidylglycine alpha-amidating monooxygenase, or PAM. The present study focused on (i) verifying that PAM could utilize a physiologically relevant peptide substrate, and (ii) demonstrating the retention of the cofactor requirements with purification of PAM. PAM (Mr = 50,000) was partially purified from rat anterior pituitary secretory granules and was shown to be capable of converting alpha-N-acetyl-ACTH(1-14) to alpha-N-acetyl-ACTH(1-13)NH2 (alpha-melanocyte stimulating hormone) and ACTH(9-14) to ACTH(9-13)NH2. The optimal rates for these conversions were dependent on ascorbic acid and CuSO4. Kinetic analyses, using the model compound D-Tyr-Val-Gly as the peptide substrate, demonstrated that, compared to the crude granule extract, the partially purified enzyme displayed increased apparent affinities for both the peptide substrate and ascorbate. These analyses also showed that the Km for D-Tyr-Val-Gly was dependent on the concentration of ascorbate, while the Km for ascorbate was constant over a wide range of D-Tyr-Val-Gly concentrations. The results presented here indicate that PAM can alpha-amidate physiologically relevant peptides related to alpha MSH, and performs the reaction in an ascorbate-dependent fashion. Retention of the ascorbate and copper requirements with purification further support the hypothesis that these cofactors are important requirements for the COOH-terminal alpha-amidation of neuro and endocrine peptides.

Adrenocorticotropic Hormone↗

PubMed1985

PTT.119, p-F-Phe-m-bis-(2-chloroethyl)amino-L-Phe-Met ethoxy HCl, a new chemotherapeutic agent active against drug-resistant tumor cell lines.

PTT.119 [p-F-Phe-m-bis-(2-chloroethyl)amino-L-Phe-Met ethoxy HCl], a new synthetic tripeptide, was highly effective against the L-phenylalanine mustard (L-PAM) resistant (L1210/L-PAM and P388/L-PAM) tumor lines, as well as the sensitive L1210 leukemia. Cytolytic activity of PTT.119 against all three leukemias was significantly greater than equimolar doses of L-PAM. These in vitro results paralleled the significant increases in mean survival times of hosts and, in some cases, abrogations of tumor formation observed in the in vivo bioassays of PTT.119-treated L1210 and L1210/L-PAM cells. Dose-response studies failed to demonstrate cross-resistance to the tripeptide by L-PAM resistant cells. Doses of PTT.119 required to reduce the viable fraction by 50% (tissue culture dose 50, TCD50) or 100% (TCD100) were 1.3- to 3-fold lower for the L-PAM resistant cells than for the L1210 leukemia. In comparison, L-PAM was unable to completely eliminate cell survival; 0.2 to 3% of the cells in all three leukemias remained viable even at doses of 75 and 163 microM. In similar studies, L1210 leukemia cells made resistant to methotrexate (L1210 MTX) and cisplatin (L1210DDP) were also completely susceptible to PTT.119; TCD50 values of the two resistant lines were 1.94 microM for L1210 MTX and 0.525 microM for L1210DDP compared to 2.38 microM for the susceptible parent L1210S leukemia. Continuous low-dose PTT.119 treatment of MJY-alpha mammary tumor cells for 8 months and exposure of L1210 leukemia to escalating levels of tripeptide for over 100 passages failed to select or induce drug-resistant phenotypes in either cell line. PTT.119 appears to be a poor mutagen and is unlikely to readily increase the probability of drug-resistant mutants in the tumor cell populations.

Animals↗