Immunoscintigraphic detection of primary and metastatic spontaneous canine osteosarcoma with F(ab')2 fragments of osteosarcoma-associated monoclonal antibody TP-1.

Monoclonal antibody TP-1 has been shown to bind selectively to human and canine osteosarcoma cells in vitro using immunohistochemical stains. This report describes the in vivo administration of radioiodinated F(ab')2 fragments of monoclonal antibody TP-1 in dogs with primary and/or metastatic spontaneous osteosarcoma. Two dogs were injected with 131labeled F(ab')2 TP-1 and two dogs were injected with 123labeled antibody fragments. Immunoscintigraphy successfully demonstrated the radiolabeled antibody fragments in 6/6 known primary or metastatic lesions and in addition detected 4 metastatic lesions not diagnosed by conventional radiographs. Concurrent imaging of 99mTc labeled autologous erythrocytes in two dogs confirmed that the accumulation of radiolabeled antibody fragments was independent of the blood pool. There was no immunoscintigraphic evidence of localization of radioiodine to normal organs or tissues other than those expected to accumulate free iodine. The present study has demonstrated the potential of monoclonal antibody TP-1 F(ab')2 fragments for early detection of metastatic spread of spontaneous osteosarcoma.


Sequence-dependent activity of 5-fluorouracil plus tauromustine in a transplantable well-differentiated murine colon adenocarcinoma.

Previous in vitro studies with 5-Fluorouracil (5-FU) plus Tauromustine (TCNU) demonstrated sequence-dependent effects. This study extended these investigations into an in vivo experimental model, relevant to clinical disease. A transplantable murine adenocarcinoma of the colon, MAC 29, was shown to have stable histology and reproducible growth. This model was used to determine the influence of sequence of administration of this drug combination. Effects ranged from antagonism, with 5-FU given 24 hours before TCNU, to addition, simultaneous treatment, and synergism, 5-FU 24 hours after TCNU. Careful consideration of sequence should be made during the evaluation of this combination in clinical trials.


Further evidence for different isoenzymic forms of a cell surface protease, guanidinobenzoatase, associated with tumours.

Normal colonic epithelial cells possess a cell surface protease referred to as guanidinobenzoatase (GB) and a corresponding cytoplasmic protein inhibitor of GB. Colonic carcinoma cells possess two isoenzymic forms of GB, the normal and the carcinoma specific form, each of which is recognised by the corresponding inhibitors present in the cytoplasm of colonic carcinoma cells. An affinity-purified inhibitor preparation obtained from the cytoplasm of cultured colonic carcinoma cells inhibited these two colonic carcinoma isoenzymic forms of GB but not the GB associated with other forms of tumour. The data suggest that each cell type possessing isoenzymic forms of cell surface GB also possesses the corresponding cell-specific inhibitors of GB.


The expression of the ras p21 oncoprotein in the bone marrow smears of children with acute leukemia.

Point mutations of the ras genes have been detected in various hematologic malignancies. This genetic event may either occur in all malignant cells or be acquired by different subclones, which however, cannot be demonstrated adequately by analyzing only DNA derived from patient specimens. The availability of the ras p21 monoclonal antibody (MoAb) Y 13259 makes possible the direct study of the distribution of the ras gene product in human malignant cells. In this report the expression of the ras p21 oncoprotein in the bone marrow smears of 35 children with acute leukemia has been analyzed. The smears were treated with the MoAb Y 13259, biotinylated goat anti-rat IgG, streptavidin, peroxidase and stained with diaminobenzidine (DAB). The intensity of the staining was evaluated by two independent observers as negative or equivocal (-/+), moderate (+) or intense (++), by counting one thousand cells. Patients were also classified according to the percentage of the stained cells into four groups (0, I, II, III). It was found that 22/35 (63%) were (+) or (++) positive as follows: 11/21 (52%) with ALL CALLA (+), 2/2 ALL-B, 3/3 ALL-T and 6/9 AML. In Group 0 (none of the blasts was stained) were 13/35 (37%), as well as in Group I (1 to 25% of the blasts stained 1+ or 2+ positive), while in Group II (26 to 50% positive stained) 3/35 and in Group III (more than 51% stained) 6/35, all of which were AML (6/9). It is concluded that the immunohistochemical analysis of the ras p21 in blast cells of children with acute leukemia may demonstrate that ras gene expression in some subclones, the intensity and percentage of which may be of some clinical importance.


Intracellular accumulation of free calcium in mouse tumor cells exposed to anticancer drugs.

Alterations in concentrations of intracellular free calcium ([Ca2+]i) were examined in mouse sarcoma S-180 cells exposed to cisplatin, mitomycin C and adriamycin, as related to cell viability. The [Ca2+]i was measured using the fluorescent Ca2+ indicator, fura-2. During the first 24 hr of incubation of the cells in vitro, in the presence of the drugs, the [Ca2+]i was lower than in the control cells. After 24 hr of incubation, the [Ca2+]i began to increase at a time when the cell viability was reduced to 50% of the controls, and finally exceeded 1 microM with reduction to 10%. The increase in [Ca2+]i was reciprocal to the loss of the cell viability. Changes in Ca2+ concentrations in the medium did not alter the results. These findings suggest that in the presence of anticancer drugs there is an accumulation of free Ca2+ with a release of this ion from internal stores and that cell death follows disruption in the homeostasis of intracellular Ca2+.


Suppressive activity of acivicin on murine bone marrow hemopoietic progenitors.

Acivicin (AVC), a L-glutamine antagonist, is an intriguing antimetabolite coupling cell growth inhibition activity with differentiating effects. In this in vivo study the influence of acivicin on mice bone marrow hemopoietic progenitors was tested. 10 mg/kg b.w./day of acivicin were i.p. injected in B6D2F1 mice for nine days. Leucocyte and reticulocyte level (in peripheral blood), CFU-S (multipotent stem cells) and GM-CFU (granulocyte-macrophage committed progenitors) content in bone marrow were determined during drug administration and for 14 days thereafter. All tested populations decreased severely during the first days of treatment. The drop was particularly striking for bone marrow CFU-S. The recovery of hemopoietic progenitors, however, began while AVC was still administered. These results suggest that the effects of acivicin on normal mouse hemopoietic system are mainly inhibitory, causing considerable myelosuppression.


Antitumor activity of orally administered SMANCS, a polymer-conjugated protein drug, in mice bearing various murine tumors.

The antitumor activity of an oily formulation of SMANCS (oily SMANCS), which is a product of conjugation between a proteinaceous antitumor antibiotic neocarzinostatin and poly (styrene-co-maleic acid), after oral administration to mice inoculated with various murine tumors was investigated. BALB/c mice, inoculated either s.c. or i.p. with allogeneic (sarcoma-180) or syngeneic (RL male 1 leukemia and Meth A fibrosarcoma) tumors, were treated with oily SMANCS orally or with an aqueous formulation of SMANCS (aqueous SMANCS) i.v. or i.p. Oral administrations of oily SMANCS or i.v. administrations of aqueous SMANCS to mice bearing three types of tumors in the ascites form resulted in a weak inhibition of tumor growth as compared to the complete inhibition of these tumors by aqueous SMANCS administered i.p. In mice bearing solid tumors, tumor growth was inhibited by 63-82% when a 10 mg/kg dose of oily SMANCS was administered orally to these mice. The antitumor potential of oily SMANCS administered orally was comparable to that obtained from solid tumor-bearing mice receiving i.v. doses of aqueous SMANCS. These results suggest that the oral administration of oily SMANCS to mice bearing various solid tumors inhibits the tumor growth as effectively as aqueous SMANCS administered i.v.


Flow cytometric DNA measurement in benign and malignant human thyroid tissues.

The occurrence of aneuploidy in 108 normal, benign and malignant thyroid tissues was evaluated by DNA flow cytometry. Aneuploid cell populations were found in 60% of the papillary carcinomas, 67% of the follicular carcinomas, 52% of the follicular adenomas and in 10% of the multinodular goiters. None of the non-neoplastic thyroid lesions (chronic thyroiditis and Graves' disease) showed aneuploidy. No significant differences in frequency of aneuploidy and DNA index were found between the well-differentiated and trabecular types of the papillary carcinomas (53%; 1.29 +/- 0.10 vs 75%; 1.30 +/- 0.10, respectively), although the latter type of papillary carcinomas have been shown to be associated with a more aggressive clinical behavior. Also no differences in frequency of aneuploidy were noted between the follicular carcinomas and the follicular adenomas, but the DNA index was significantly higher in the former. These findings indicate that flow cytometric analysis of DNA content may not be helpful in the evaluation of the clinical behavior of papillary carcinomas or in the determination of malignancy of follicular tumors, but may be useful in differentiating neoplastic from non-neoplastic thyroid lesions.


Mutant p53 product in patients with stage III cervical cancer.

Mutations in the p53 gene stimulate cell division. In our study we assessed the prognostic value of mutant p53 overexpression in cervical cancer stage FIGO III detected by immunohistochemistry. In 43 tissue specimens were detected p53 overexpression in 20 cases. Mean survival time was 36.4 (SE +/- 7.66) months in the p53 protein positive group. The group without p53 overexpression showed a mean survival time of 28.6 (SE +/- 3.85) months. p53 protein overexpression had no prognostic value in patients with stage III cervical cancer.


Total control of chemotherapy induced emesis.

A combination of lorazepam, prochlorperazine, dexamethasone, scopolamine and ondansetron was used to prevent and diminish the occurrence of nausea and vomiting during administration of emetogenic chemotherapy. Immediately prior to administering chemotherapy a cutaneous scopolamine patch was applied to the retroaural area, prochlorperazine 5 mg was given orally, and intravenous (IV) lorazepam 0.5 mg, ondansetron 10 mg and dexamethasone 10 mg were given. The last two were repeated after 4 hours twice, and lorazepam at 6 hour intervals for four doses. A chemotherapy course consisted of mitoxantrone 20 mg/m2 i.v., followed by thiotepa 60 mg/m2 followed by 3 grams/m2 of cyclophosphamide as a constant infusion over 48 hours. There were 80 courses given to 30 women and one man with a median age of 42 years (29-58). Complete protection from vomiting in all 80 courses resulted. Minor nausea occurred in 25 courses. The antiemetic regimen was well tolerated and all patients received all the doses as prescribed.


Production of human macrophages with potent antitumor properties (MAK) by culture of monocytes in the presence of GM-CSF and 1,25-dihydroxy vitamin D3.

Antitumoral macrophages (MAK) were obtained by the culture of human mononuclear cells in hydrophobic bags. From one cytapheresis, up to 10(9) mature macrophages could be purified by elutriation after one week of culture in IMDM medium in the presence of 2% human AB serum. These MAK cells were used for adoptive treatment in metastatic cancer patient with no dose-limiting toxicity. The present study aimed to improve the average MAK yield by addition of GM-CSF and of dihydroxy-cholecalciferol. The differentiated macrophages obtained presented higher antitumoral functionality in response to rh-IFN gamma than in their absence. These MAK presented all the differentiation antigens of cytotoxic macrophages compared to MAK cells differentiated in standard medium. They killed human tumor targets effectively in vitro at a low (1/1) effector/tumor ratio; furthermore, the antitumoral activity reached by MAK cells after IFN gamma activation appeared to be stabilized for several days.


Mechanism of action of tricyclic drugs on Escherichia coli and Yersinia enterocolitica plasmid maintenance and replication.

Tricyclic medical compounds like many other non-antibiotics exhibit antimicrobial activities. Two chemically representative groups were tested in plasmid DNA transformation and replication to assign intracellular target sites responsible for the multiple effects in Escherichia coli and Yersinia enterocolitica cells. To analyse the mechanism of action at the molecular level, the effects of chlorpromazine, 7,8 dioxochlorpromazine, promethazine, methylene blue, imipramine, cannabidiolic acid and tetrahydrocannabidiolic acid were examined at several points in the course of transformation, in plasmid replication and on the topological state of plasmid DNA. Two possible target sites were identified, one of them involving membrane binding sites which participate in plasmid DNA replication. Drug binding at these sites interfered with the replicating plasmid DNA and membrane protein complex, preventing the proper processing of the replication that resulted in plasmid loss. The other in vivo and in vitro effect was observed on the topological state of plasmid DNA. Tricyclic drugs intefered with energy dependent gyrase activity and promoted the relaxation of plasmid DNA, causing disturbances in gene expression and in plasmid replication. The results give insight into the chemical structures connected with significant specific antimicrobial effects.


Ly-6A/E gene is widely expressed among transformed nonhematopoietic cells. Autocrine modulation by interferon.

Expression of the Ly-6A/E gene by transformed cells was investigated in 14 cell cultures of C57BL/6 and BALB/c mouse origin derived from spontaneous or chemically-induced non-hematopoietic tumours and from cells transformed by SV40 virus. Histologic types included carcinomas, sarcomas and a melanoma. Thirteen out of 14 cell cultures expressed membrane Ly-6A/E antigens; only the B16-A melanoma was negative, but it expressed Ly-6A/E after incubation with IFN-gamma. The effect of in vitro permanence was studied on early (< 10) and late (> 20) passages of B16-A melanoma and MN/MCA1 fibrosarcoma. Late passage B16-A cells were Ly-6A/E-negative and refractory to induction of Ly-6A/E (but not of H-2 antigens) by IFN-alpha, IFN-beta, or IFN-gamma; MN/MCA1 maintained a high expression of Ly-6A/E, but no increase was induced by IFNs. It was found that both early and late in vitro passages of MN/MCA1 actively produced IFN-alpha/beta. The analysis of cells of fibroblastic origin revealed a significant correlation between IFN release in the culture medium and Ly-6A/E levels. Culture of fibrosarcoma cells in the presence of an anti-IFN-alpha/beta serum reduced Ly-6A/E expression, thus indicating the existence of an autocrine loop.


Characterization of nuclear size, ploidy, DNA histogram type and proliferation index in 79 nerve sheath tumors.

The distribution values of ploidy, of the DNA histogram type, of the proliferation index and of the nuclear area are described in a series of 79 nerve sheath tumors including 7 traumatic neuromas, 55 primary and 9 recurrent benign schwannomas, 5 benign neurofibromas and 3 malignant tumors. The nuclear parameters were computed on Feulgen-stained nuclei (from archival formalin-fixed paraffin-embedded materials) by means of digital cell image analysis. The results show that 20-30% of benign nerve sheath tumors exhibit an abnormal nuclear DNA content (DNA index > 1.30). Such a feature was observed in the four groups of benign tumors, i.e. primary and recurrent schwannomas, neurofibromas and even traumatic neuromas. Aneuploid tumors exhibited triploid, hypertriploid or polymorphic DNA histogram types. The three malignant tumors under study were aneuploid. Lastly, the nuclear area and proliferation index measurements did not allow any clear distinction to be made between the four groups of benign tumors on the one hand, and between the benign and malignant tumors on the other.


Synthesis and antitumor activity of a new cis-diammineplatinum (II) complex containing procaine hydrochloride.

This paper refers to some of the chemical and biological properties of a new platinum (II) complex where the aromatic amino group of procaine is involved in the coordination with platinum and whose structure was defined by UV, IR, 1H-NMR, and elemental analysis. This new cationic platinum-triamine complex (DPR) displays excellent solubility (> 50 mg/ml) and stability in water. DPR has significant in vitro cytotoxicity against murine P388 leukemic cell line, human K562 erythroleukemic cell line and human Jurkat T cell line. The in vitro cytotoxic effects of DPR on P388 and Jurkat leukemic cells were comparable to those of cis-diamminedichloroplatinum (II) (DDP), while its activity on K562 cells was significantly better than that of DDP [IC50 = 1.07 +/- 0.36 (SD) microM vs 2.62 +/- 0.23 (SD) microM, P < 0.01]. The in vitro Pt accumulation rate for P388 cells was twice as rapid after DPR than after DDP exposure, while no difference in cellular platinum efflux was observed. The antitumor activity of DPR was tested in vivo against P388 leukemic cells in BDF1 mice and gave a % ILS value (75%) similar to that of the maximum tolerated dose (MTD) of DDP (8 mg/Kg). A comparative study of plasma urea nitrogen (PUN) levels and kidney morphological analysis in tumor-bearing mice receiving the LD50 dose of both drugs (39.3 mg/Kg and 16.5 mg/Kg for DPR and DDP, respectively), showed DPR to be less nephrotoxic than DDP. These results indicate that this new cationic platinum-triamine complex containing primary amine ligand is surprisingly active both in vitro and in vivo. In summary, the good characteristics of DPR in terms of high solubility, encouraging anticancer activity and absence of nephrotoxic effects make DPR a promising new platinum anticancer agent for preclinical development.


Tissue-specific induction of metallothionein by bismuth as a promising protocol for chemotherapy with repeated administration of cis-diamminedichloroplatinum (II) against bladder tumor.

The effects of bismuth nitrate pretreatment on the toxicity and antitumor activity of repeatedly administered cis-diamminedichloroplatinum (Cisplatin; CDDP) were examined using nude mice inoculated with human bladder tumor tissues. Lethal and renal toxicities exerted by the repeated administration of CDDP were effectively prevented by pretreatment with bismuth (Bi) without affecting its antitumor activity against transplanted human bladder tumor as in the case of single dose of the drug reported previously. The renal Bi level was gradually increased with the frequency of Bi administration, and metallothionein (MT) induced by Bi in the kidneys maintained its substantially high level during the treatment. It was confirmed that MT was not induced in the tumors even by the 5 cycles of repeated Bi administration. This specific protection shown by the Bi preadministration against the toxicity of repeatedly injected CDDP can be explained by the fact that Bi markedly induces MT in the kidney, a major target organ of CDDP toxicity, but not in the tumor tissues inoculated in the nude mice, probably because Bi is efficiently taken up by the kidney but hardly incorporated into the tumor tissues as reported previously. These data obtained by repeated doses of CDDP as described above strongly suggest a promising protocol for chemotherapy using CDDP with Bi compounds, a tissue specific MT inducer, against advanced bladder tumor in human.


A prospective study of the value of imaging, serum markers and their combination in the diagnosis of pancreatic carcinoma in symptomatic patients.

The diagnostic accuracy of ultrasound (US), computed tomography (CT), endoscopic retrograde cholangiopancreatography (ERCP), and tumour markers CEA, CA 50 and CA 242 in pancreatic cancer (n = 26) was studied in 113 patients with jaundice, in 20 patients with unjaundiced cholestasis, and in 60 patients with the suspicion of chronic pancreatitis or a pancreatic tumour. The sensitivities of US, CT and ERCP were 61.9%, 95.2% and 82.3%, the specificities 93.9%, 92.9% and 94.1%, and the efficiencies 91.6%, 96.6% and 92.1%, respectively. The sensitivities of CEA, CA 50 and CA 242 were 92.3%, 96.1% and 61.5%, the specificities 59.2%, 58.0% and 95.2%, and the efficiencies 63.7%, 63.2% and 90.6% respectively. The combined use of the imaging methods and tumour markers was also analysed. When either the imaging method or the serum marker test was required to be positive, the sensitivities of the combinations were clearly better than those of US and CA 242 alone, but only slightly better than those of CT, ERCP or the tumour markers CEA and CA 50 alone. When both the imaging test and the marker test were required to be positive, the specificities of the combinations were clearly better than those of CEA and CA 50 alone, but they did not exceed the specificity of the imaging methods or CA 242 alone. We conclude that CT, ERCP and CEA and CA 50 are highly sensitive in the diagnosis of pancreatic cancer in symptomatic patients, while the sensitivity of US and CA 242 is lower. The specificity of the imaging methods and CA 242 is high, but that of CEA and CA 50 is low. Imaging methods and serum tumour markers could be more used in clinical practice in a complementary manner. In patients with jaundice and/or cholestasis or with a suspicion of pancreatic tumour or chronic pancreatitis, the combined use may yield higher sensitivity than US alone and higher specificity than CEA or CA 50 alone.


Studies of the digestibility of pigs fed dietary sucrose, fructose or glucose.

Digestion of sucrose, fructose and glucose was studied in pigs with reentrant cannulas in the distal ileum or intact pigs fed diets formulated with sucrose, fructose and glucose as the major energy source and torula yeast. Sucrose and glucose were completely digested in the small intestine (apparent digestibility, 98.3 and 98.3%, respectively but a fraction of the dietary fructose escaped to the large intestine (adjusted value, 11.7%). There was no carbohydrate in rectal contents of the pigs. There was no carbohydrate influence on the pre-caecal or total digestibility of nitrogen.


[The extraction particles from winter rapeseed with different glucosinolate fractions in swine with regard to the iodine supply. 3. Apparent digestibility of crude nutrients with special regard to the carbohydrate and nitrogen balance].

In two digestibility and N-balance experiments the following solvent extracted meals were examined in a grain (barley and wheat in equal parts) or in a starch diet with four female pigs/group fitted with urethra catheters during the 5 days sampling period: 48% soyabean meal (SBM) (1), 24% conventional rapeseed meal (RSM) (2), 24 (3) or 48% (4) RSM from a newly bred low glucosinolate variety, groups 1 to 4 with grain, 48% low glucosinolate RSM (5) or 48% SBM (6) both with starch. In numeric order of the groups 88, 85, 86, 84, 79 and 89% N were digested. Pigs responded to the additional faecal N excretion from RSM diets by a lower urinary N excretion and thus they reached the N-balance of the animals fed on SBM. In numeric order of the groups 86, 79, 80, 73, 73 and 92% of the organic matter of the solvent extracted meals were digested. In comparison with SBM even 48% RSM in the feed significantly lowered the digestibility of organic matter. Using detergents the additional faecal organic matter excretion of RSM versus SBM could quantified as two third lignin. In the case of 24% RSM in the diet--i.e. 15% of the carbohydrates given--the carbohydrate digestibility is estimated with a high error, that an energetic feed value should not calculated. Investigating 48% SBM or RSM respectively in a grain diet the net energy content was 9.8 MJ (666 Energetic Feed Units, EFU pig) or 8.3 MJ (569 EFU pig)/kg dry matter (DM). Based on the starch diet, however, little relevant for estimation of normal pig diets, there were evaluated 10.6 MJ (720 EFU pig) for SBM and 8.3 MJ (567 EFU pig) net energy/kg DM for RSM.


[Effect of the manganese content in laying hen feed with different Ca and mineral levels on the egg shell quality and bone mineralization of hens].

Four experiments with 270, 44, 432 and 66800 Leghorn hens were carried out to investigate the influence of various Mn additions to diets differed in mineral or Ca contents on egg shell quality. The addition of 300 mg Mn/kg diet improved significantly egg shell breaking strength by 4 N over one year. The supply of 50-500 mg Mn/kg diet for 10-24 weeks of the second half of laying year did not influence the egg shell quality. Addition of mineral mixture or Ca grit to layer rations with adequate or higher Mn levels did not influence egg shell strength. High mineral content in a low manganese diet increased number of cracks by 3%. Strength, weight and ash content of tibia were significantly reduced by feeding a low mineral level. Addition of 50-150 mg Mn per kg low mineral diet normalized partially tibia stability in young hens. It was concluded that supplied dietary Manganese influences calcification positively only in young hens. High levels of Ca did not influence the effects of Mn. 50 mg Mn per kg layers mixture have been considered as an essential supply.


Uncommon mesenchymal tumors of the liver. Light microscopy, ultrastructure, immunohistochemistry and literature review.

This report presents the gross morphology, cytological, histological, ultrastructural and immunohistochemical features of two angiomyelolipomas of the liver. Diagnosis and classification of these tumors is difficult. There is no immunohistochemical and ultrastructural confirmation for Ito-cell pathogenesis of hepatic angiomyelolipomas.


DAP-decarboxylase activity and lysine production by rumen bacteria.

The last step of pathway of lysine biosynthesis by rumen bacteria was tested. The first measurements of DAP-decarboxylase activity and of lysine production by Megasphera elsdenii, Selenomonas ruminantium, Clostridium spp., Butyrivibrio fibrisolvens and Bacteroides succinogenes as well as the first attempts to increase the lysine production by ruminal streptococci by mutation are described. The highest values were measured in Selenomonas ruminantium (DAP-decarboxylase activity = 146 micrograms DAP.min-1.mg-1 protein and lysine production was 390 micrograms.mg-1 protein) and the lowest values were ascertained in Butyrivibrio fibrisolvens (DAP-decarboxylase activity = 27 micrograms DAP.min-1.mg-1 protein and lysine production was 32 micrograms.mg-1 protein). DAP-decarboxylase activity was increased by mutation especially in Streptococcus bovis, the lysine production in both of tested ruminal streptococci. The potential use of lysine-excreting mutants in calves in future is suggested.


Immunohistochemical detection of mutant p53-suppressor gene product in patients with breast cancer: influence on metastasis-free survival.

Overexpression of p53-protein appears to be a common event in primary breast cancer. It has been proposed that the presence of elevated levels of this protein may be an independent prognostic factor and may be important for the ability of a tumor to metastasize. This study was performed to evaluate the influence of immunohistochemically detectable mutant p53-protein on metastasis-free survival of patients with breast cancer. Immunohistochemistry was performed on 117 paraffin-embedded biopsy specimens of consecutive patients with stage T1-T4 breast cancer, using a monoclonal antibody against p53 suppressor gene product. 29 (24.8%) specimens showed positive staining, whereas in 88 (75.2%) a negative staining reaction for p53 was found. Comparing time intervals to diagnosis of metastasis, using Kaplan-Meier curves, Log-Rank test revealed no significant differences in metastasis-free survival between p53 positive and negative patients (P = 0.32), whereas statistically significant differences were noted for tumor stage (P < 0.01), nodal status (P < 0.01), histological grading (P < 0.01) and estrogen receptor status (P = 0.03). Mutant p53-protein, as detected by immunohistochemistry in paraffin embedded tumor tissue, does not appear to influence metastasis-free survival in patients with breast cancer.


Digestion and performance responses to lasalocid and concentrate supplements by beef cattle fed bermudagrass hay.

Beef cattle consuming bermudagrass hay were not supplemented or received a limited amount of ground corn alone or with a mix of protein meals to determine influences of concentrate supplementation on digestion and performance when the ionophore lasalocid (200 mg daily) was given. With limited feed intake, supplement treatment did not change the acetate to propionate shift in beef cows occurring with lasalocid (P < 0.06). Lasalocid did not affect sites of digestion of organic matter or nitrogen with any supplement treatment. However, lasalocid decreased (P < 0.10) ruminal digestion of neutral and acid detergent fibre. Live-weight gain by growing beef calves ingesting bermudagrass hay ad libitum was higher (P < 0.05) with than without supplementation and tended (P < 0.12) to be greater for corn plus protein meals than for corn alone. Lasalocid did not affect or interact with supplement treatment in feed intake or live-weight gain of heifers (236 kg; no growth stimulant) or steers (237 kg; treated with 200 mg progesterone and 20 mg estradiol benzoate). Lasalocid at 200 mg daily did not improve digestion characteristics or influence performance by beef cattle consuming a Basal diet of bermudagrass hay. Further, effects of lasalocid were not modulated by supplementation with concentrate, concentrate type or sex or growth stimulant usage.


Temperature- and concentration-dependence of compatibility of the organic osmolyte beta-dimethylsulfoniopropionate.

The effects of the organic osmolyte beta-dimethylsulfoniopropionate (DMSP) on the structural stability of three model proteins were examined to determine whether DMSP, like the structurally similar solute dimethyl sulfoxide (DMSO), is compatible with native protein structure at low, but not elevated, temperatures. DMSP stabilized phosphofructokinase under conditions of cold-induced denaturation. Thus, DMSP, like DMSO, may be an effective protein cryoprotectant. However, DMSP was not an effective stabilizer of protein structure under conditions of heat denaturation. Whereas low (0.2 M) concentrations of DMSP stabilized lactate dehydrogenase against inactivation at 50 degrees C, higher DMSP concentrations were ineffective. DMSP favored the denaturation of glutamate dehydrogenase at all DMSP concentrations tested. DMSP may be a compatible osmotic solute only under conditions of moderate temperature and low, yet physiological, concentrations. The mechanistic basis of DMSP's temperature- and concentration-dependent effects and the possible roles played by adaptation temperature and severity of osmotic stress in the evolutionary selection of organic osmolytes are discussed.


Photolabelling of Salmonella typhimurium LT2 sialidase. Identification of a peptide with a predicted structural similarity to the active sites of influenza-virus sialidases.

The sialidase from Salmonella typhimurium LT2 was characterized by using photoaffinity-labelling techniques. The well-known sialidase inhibitor 5-acetamido-2,6-anhydro-3,5-dideoxy-D-glycero-D-galacto-non- 2-enonic acid (Neu5Ac2en) was modified to contain an amino group at C-9, which permitted the incorporation of 4-azidosalicylic acid in amide linkage at this position. Labelling of the purified protein with the radioactive (125I) photoprobe was determined to be highly specific for a region within the active-site cavity. This conclusion was based on the observation that the competitive inhibitor Neu5Ac2en in the photolysis mixture prevented labelling of the protein. In contrast, compounds with structural and chemical features similar to the probe and Neu5Ac2en, but which were not competitive enzyme inhibitors, did not affect the photolabelling of the protein. The peptide interacting with the probe was identified by CNBr treatment of the labelled protein, followed by N-terminal sequence analysis. Inspection of the primary structure of the protein, predicted from the cloned structural gene for the sialidase [Hoyer, Hamilton, Steenbergen & Vimr (1992) Mol. Microbiol. 6, 873-884] revealed that the label was incorporated into a 9.6 kDa fragment situated within the terminal third of the molecule near the C-terminal end. Secondary-structural predictions using the Garnier-Robson algorithm [Garnier, Osguthorpe & Robson (1978) J. Mol. Biol. 120, 97-120] of the labelled peptide revealed a structural similarity to the active site of influenza-A- and Sendai-HN-virus sialidases with a repetitive series of alternating beta-sheets connected with loops.


Pulsatile flow visualization in the abdominal aorta under differing physiologic conditions: implications for increased susceptibility to atherosclerosis.

The infrarenal abdominal aorta is a common site for clinically significant atherosclerosis. As has been shown in other susceptible locations, vessel geometry, flow division rates, and pulsatility may result in hemodynamic conditions which influence the preferential localization of disease in the abdominal aorta segment. Pulsatile flow visualization was performed in a glass model of the aorta constructed from measurements of angiograms and cadaver aortas. Flow rates and pulsatile waveforms were varied to reflect typical physiological conditions. Under normal resting conditions, the flow patterns in the infrarenal aorta were more complex than those in the suprarenal location. Time varying vortex patterns appeared at the level of the renal arteries and propagated through the infrarenal aorta into the common iliac arteries. A region of oscillating velocity direction extended from the renal arteries to the aortic bifurcation along the posterior wall. Dye became trapped along the posterior wall, requiring several cardiac cycles for clearance. In contrast, there was rapid clearance of the dye in the anterior aorta. Under postprandial conditions, the flow patterns in the aorta were basically unchanged. Simulated exercise conditions created laminar hemodynamic features very different from the resting conditions, including a decrease in dye residence time. This study reveals significant time-dependent variations in the hemodynamics of the abdominal aorta under differing physiologic conditions. Hemodynamic factors such as low wall shear stress, oscillating shear direction, and high particle residence time may be related to the clinically seen preferential plaque localization in the infrarenal aorta.


Cage allocation designs for rodent carcinogenicity experiments.

Cage allocation designs for rodent carcinogenicity experiments are discussed and presented with the goal of avoiding dosage group biases related to cage location. Considerations in selecting a cage design are first discussed in general terms. Specific designs are presented for use in experiments involving three, four, and five dose groups and with one, four, and five rodents per cage. Priorities for balancing treatment groups include horizontal position on shelf and shelf of rack, nearest neighbor balance, and male-female balance. It is proposed that these balance criteria be considered together with practical issues, such as the ability to accurately conform to a design and to determine a sensible and efficient design for each experiment.


Estimating gallstone incidence from prevalence data.

Because incidence data for gallstones are generally unavailable, we have used a previously described epidemiologic method to estimate age-specific incidence rates from published gallstone prevalence data in seven populations. Yearly incidence rates ranged from a low of 1 in 1000 for the younger men to 19 in 1000 for the oldest women. An overall pattern of increasing incidence of disease with increasing age was observed. Rate-ratios for oldest versus youngest subjects ranged from 4.0 in German women to 9.4 in Italian men: mean rate-ratio = 5.9; 95% CI = 4.0-7.8. For all centers, age-specific female incidence rates exceeded comparable age-specific male rates at all ages (mean female to male rate-ratio = 1.85; 95% CI = 1.7-2.0). No differences were observed in the sex-specific incidence of gallstones for subjects of similar age in the various centers (P = > 0.10). For Denmark our estimates resemble recently published incidence data obtained by re-surveying the original population.


[Role of home monitoring in the context of sudden infant death prevention. Current methods indications, monitoring].

Some infants are cared with a home monitoring system during their first year of life. An international clinical consensus has been obtained and has proposed this technique mainly for infants who have presented an apparent life threatening event or for ex-premature with bradycardia or apnea, rather than for siblings of sudden infant death syndrome or other infants. In any case, this monitoring must be held after a complete clinical evaluation of the infant and after a real education of the parents about the use of the device. Many types of devices are used. The most efficient is the cardio-respiratory monitoring. Some of them include a processor and record the alarms. The need to see or to call the medical team to decode them allows close collaboration between the family and the clinical team. Knowledge of the alarms and the circumstances in which they have occurred help the medical team to propose the withdrawal of the home monitoring. Thus, sometimes preventive, sometimes prophylactic, this device will provide us for an optimal help.


Carotid endarterectomy in Puerto Rico.

The experience of the authors performing carotid endarterectomy in Puerto Rico is reported. The study was stimulated by the recently published results of the Carotid Endarterectomy Cooperative Trial groups in North America and Europe. This series consists of 61 carotid endarterectomies performed on 53 patients. The majority of the patients suffered from hypertension, diabetes mellitus, smoking, and ischemic heart disease. Most of the patients presented with Transient Ischemic Attacks (64%) or Reversible Ischemic Neurologic Deficits (19%). One patient died of a presumptive myocardial infarction and one patient had a post-operative worsening of his neurologic condition. The permanent morbidity and mortality rate was 3.2%.


Usefulness of single fibrinopeptide A determination in patients with acute ischemic coronary artery syndromes.

Coronary artery thrombosis plays a major role in the acute ischemic coronary artery syndromes in which fibrinopeptide A (FPA) has proved to be a sensitive marker. The purpose of this study was to determine FPA concentrations in patients with acute coronary artery syndromes and to determine if these could serve as a short-term prognostic indicator. Single plasma FPA levels were measured in 26 patients with acute ischemic coronary artery syndromes within 24 hours of the onset of chest pain as well as in 12 patients with chronic stable angina and in 9 control subjects. Higher FPA levels were observed in patients with unstable angina whom later developed recurrence of chest pain compared to those without (8.1 +/- 3.4 vs. 3.4 +/- 2.2; p = 0.01). Neither the localization of ischemia, presence of complications, need for revascularization nor short-term prognosis (6 months) correlated with the plasma FPA concentration. Therefore, except for recurrence of chest pain in patients with unstable angina, the finding of an elevated FPA level upon admission did not provide additional information regarding clinical course and prognosis than that obtained in a detailed clinical history, physical examination and initial electrocardiogram in patients with acute ischemic artery syndromes.


Cardiac surgery on Jehova's Witnesses at Instituto Cardiovascular-Hospital Pavia.

Jehovah's Witnesses (J.W.) can undergo successful cardiac operations. We have operated five J.W. patients. Of these patients, two had coronary artery bypass surgery and three had correction of congenital anomalies. These included an atrial septal defect with infundibular pulmonic stenosis, a tetralogy of Fallot and a patient with a ventricular septal defect. Our treatment protocol includes a meticulous surgery, the use of early heparinization to collect all shed blood into the pump oxygenator, observation in the operating room for early exploration if the patient bleeds and administration of iron preparations. Recombinant human erythropoietin, although available and in our treatment protocol, has not been used yet. All patients survived the operation and left the hospital with an excellent hemoglobin and hematocrit. The length of stay varied from 7 to 15 days.


Hypertension: pathophysiologic-based management. Part II.

The therapy for hypertension utilizing diuretics and beta-blockers induces metabolic and biochemical alterations that blunt the possible beneficial effects of blood pressure reduction in coronary artery disease risk. The utilization of other antihypertensive medications with the capacity of improving cardiovascular risk factors, like diabetes and dyslipidemias is recommended.


Cardiopulmonary resuscitation personal choices of physicians and nurses.

Nine hundred and twenty-two attending physicians, residents and graduate nurses in five hospitals were requested to state their personal preference to receive either cardiopulmonary resuscitation (CPR) or intubation and mechanical ventilation (MV) when given three estimated outcomes: 1) complete recovery, 2) important loss in mental and physical functions, c) persistent unconsciousness. They also could choose not to accept either procedure. Seventy three percent would accept CPR and 70 percent MV, only if complete recovery was likely. Thirteen percent would accept CPR and MV if important losses of function were likely. Eight percent would accept CPR and 5 percent would accept MV if recovery of consciousness was unlikely. Six percent would not accept CPR and eleven percent would not accept MV under any circumstance. Men and women differed in the choice of outcomes which made resuscitation acceptable to them (p < .001). More women than men would accept CPR and MV if complete recovery was likely and were unwilling to accept CPR or MV under any circumstance at all. Half as many women as men would accept MV if they were likely to remain unconscious. Choices also varied with age and professional training. There are large individual differences as to how physicians and nurses would like their personal cardiopulmonary arrest to be managed. Their choice is appropriately influenced by the expected outcome and seems to be influenced by the local hospital setting.


Working with the elderly: do student nurses care for it?

This study investigates neophyte student nurses' attitudes to working with the elderly through placing them in relation to attitudes to other nursing career options and by exploring student nurses' reasons for such attitudes. The results are based on a questionnaire answered by 610 students from five NSW teaching institutions. The results indicate that nurses rate working with the elderly very poorly, and that this negativity is based on unfavourable stereotypes of aged persons and their care. These findings are placed in the context of projected demographic changes and the pressures these will place on the health care system and its need for nurses qualified in this area of work.


Child immunisation levels in Sydney's Western Metropolitan Region: parental attitudes and nurses' roles.

Research was undertaken to examine childhood immunisation uptake rates, parental attitudes towards immunisation and immunisation services and parents' perceptions of health workers' (especially nurses') contribution to immunisation in the Western Metropolitan Region of Sydney, NSW. After a pilot study a questionnaire was distributed by teachers to a random sample of 450 parents of kindergarten children living in the Region. Data revealed an 84% childhood immunisation rate for all scheduled immunisations up to five years with a partial immunisation rate of 10% (6% did not answer or had gaps in immunisation information). The most significant factors associated with partial immunisation were found to be the socioeconomic and educational status of the children's fathers and itinerancy. Parents of partially immunised children were significantly more likely than parents of totally immunised children to consider that serious side-effects to immunisation do occur. The research also found that parents have a knowledge deficit about the seriousness of measles; that immunisation services in the Region have significant deficiencies and that nurses have a low profile and poor image in the provision of those services. As Early Childhood nurses were considered by parents as providing the most appropriate immunisation nursing service, their further involvement in promoting and extending services is recommended.


Nurse practitioners' and students' attitudes towards people with disabilities.

This article reports the results of assessments of nurse practitioners' and nursing students' attitudes toward people with disabilities. Findings of three projects that used the Interaction with Disabled Persons Scale suggest that Australian nurses' and nursing students' attitudes are more positive than those of the general population and that nurse education strategies are effective in promoting positive attitudes. These findings contrast with the results of overseas studies which indicate that health professionals' attitudes toward people with disabilities are negative and unaccepting.


[Changes due to age in the regenerating axons of the adult rat].

Muscle reinnervation after nerve crush was observed in rats at different ages with a combined technique that simultaneously demonstrates nerve endings and endplates. At early times of reinnervation the amount of sprouting was higher in older rats than in younger rats; according to this finding an enhanced number of polyinnervated endplates was found in older rats. A similar enhancement of sprouting and polyinnervation was observed during muscle reinnervation of vitamin E deficient rats, supporting the proposed analogy between vitamin E deficiency and aging.


Handover: the collective narrative of nursing practice.

This paper examines the practice of handover in a large metropolitan hospital. It shows that the handover is a significant site at which to examine how tensions and imperatives derived from the traditional institutional position and role of the nurse are played out in contradiction with emergent professionalism. It identifies handover dimensions and focuses discussion on how the collective narrative of the handover serves to construct patient identities as well as ensure solidarity and cohesion among nurses.


Calcium adenosinetriphosphatase of bovine retinal rod outer segment disks.

A Ca(2+)-pumping ATPase activity is present in bovine retinal rod outer segment purified disks. The ATPase has a high Ca2+ affinity (KM = 25 microM). Low Ca2+ (n-microM) concentrations stimulate an ATP-dependent Ca2+ uptake and the ATP hydrolysis in the absence of exogenous Mg2+. Electrophoretic analysis of disk proteins after treatment with (gamma-32P)ATP shows the existence of the enzyme-phosphate acid-stable, hydroxylamine-sensitive intermediate complex of molecular mass of about 135 kDa. The results would indicate the presence of an inwardly directed Ca(2+)-ATPase pump acting on the disk membrane, that could be involved in the regulation of cytosolic free Ca2+ levels inside ROS.


Queen Elizabeth Behavioural Assessment Graphical System.

The Queen Elizabeth Behavioural Assessment Graphical Scale (QEBAGS) is presented as a new, descriptive rating method for documenting behavioural disturbances in a variety of clinical settings. The scale utilises three categories of behavioural disturbances which may occur in isolation or may co-exist. These are documented on a graphical plot across a 24 hours' time span. Neither diagnostic information nor severity of a particular behavioural disturbance needs to be interpreted before a rating on each parameter can be made, thus allowing its use by any intelligent observer, including a carer at home. Documenting of behavioural disturbance across day and night, by intermittent or regular observations, provides valuable information on the time, course and pattern of behavioural disturbance, from which diagnostic and therapeutic directions can be inferred. In preliminary clinical use, the scale has improved documentation of behavioural disturbance and communication between health professionals about such behaviour and its management. The scale also has the potential to provide supportive documentation for the Resident Classification Index in Nursing Homes and Personal Care Assessment Index in Hostels. The scale is most effective in situations where moderate to severe disturbance of behaviour is present. Where there is little or no disturbance, the system is unlikely to be of benefit. Clinical examples of its utility are presented.


[Fluorocytometric study of proliferation antigens, nuclear proteins and ploidy in acute leukosis].

P53 protein, Ki67 proliferative associated antigen and DNA content have been studied by flow cytometry in the blood blastic cells from 41 patients with acute leukemia. The results were compared with the F.A.B. classification. Cells were permeabilised and fixed by PLP solution before using the FITC conjugated Ki67 MoAb and the p53 MoAb (clone 1801). Propidium iodide and RNAase has been used in order to determine ploidy. Ki67 and p53 protein were found to be expressed at higher level in leukemia cells than in normal bone marrow cells; however there was no correlation between Ki67 and p53 expression and F.A.B. subtype. In acute leukemia patients the range of positivity of Ki67 was 1.1-52.1% while it ranged from 1.8% to 80.1% for the p53 protein. On the basis of these findings we conclude that the flow cytometry evaluation of Ki67 and p53 represents a useful tool for the study of the biologic characteristics of the leukemic cells in patients with acute leukemia.


[Effect of ASA on the interaction of von Willebrand factor with the platelet membrane].

Ristocetin induces a conformational change on von Willebrand Factor (vWF) similar to that due to the interaction with the subendothelium, by which the former can interact with the Glycoprotein-1 B (GPIB) of the platelet membrane and trigger aggregation and granule content secretion. Platelet Rich Plasma (PRP) treated with Acetyl Salicylic Acid (ASA) loses completely the aggregability induced by addition of Ristocetin whereas ASA-treated and successively Washed Platelets (AWP) supplemented with normal plasma (PPP) give an aggregation and a secretory response to Ristocetin similar to that given by PRP; similarly normal Washed Platelets (WP) supplemented with ASA-treated plasma (APPP) give identical aggregation, and secretion by Ristocetin addition. Ours results indicate that the Ristocetin-vWF complex can trigger two distinct intraplatelet metabolic pathways. A first well known way starts from the activation of Phospholipase A-2 (PL-A2), by which arachidonic acid is produced, that, in turn, undergoes the metabolic pathway leading to Thromboxane A-2; this pathway can be blocked by the intraplatelet ASA by irreversible inactivation of Cyclooxygenase, but it is insensitive to the extra-platelet ASA. A second, independent metabolic pathway, can be triggered by intact vWF, but not by the ASA treated one. It is insensitive to intraplatelet ASA and therefore unrelated to the arachidonic acid metabolism. This pathway could start from the activation of Phospholipase C (PL-C).


Ascorbate peroxidase activity in resistant and susceptible plants of Lycopersicon esculentum.

Activity of redox-enzymes of AA system and of catalase was measured in two near-isogenic tomato lines, respectively resistant and susceptible to Tobacco Mosaic Virus infection. AFR reductase, DHA reductase and catalase showed quite similar activities in both lines, whereas AA peroxidase activity in resistant plants was 75% higher than in susceptible ones, with Km values about 4-fold lower. These data suggest that hydrogen peroxide scavenging operated by AA peroxidase could play an important role in the development of biological defence mechanisms against pathogens.


Single stranded DNA-vectors for analyzing processing of DNA damage induced by 4-nitroquinoline-1-oxide in prokaryotes and eukaryotes.

Previous studies indicate that single stranded DNA vectors could be used in different organisms to study mutagenesis induced by DNA damaging agents. We applied this approach to study mutagenesis induced by 4NQO lesions. The use of ssDNA, on which the ultimate metabolite of 4NQO (Ac-4HAQO) induces mainly C8-guanine adducts, allowed us to find a correlation between G-transversions and the dGuo-C8-AQO adduct. This correlation was established in two independent assay-systems, based on prokaryotic and eukaryotic cells.


Investigation of the patterns of argyrophilic nucleolar organiser regions (AgNORs) in primary gastric lymphomas.

Eight cases of primary gastric lymphomas have been investigated by AgNORs method to individualize the patterns of distribution of NORs. Modifying and exemplifying the scheme of Nikicicz and Norback, previously applied to blood smears and bone marrow in patients affected by leukaemia, the authors found 8 principal distribution patterns of AgNORs. Recording the percentage of the single patterns in every case it was possible to individualize two quite homogeneous groups. The authors maintain that in this way, avoiding complex numerical evaluations and statistical analysis, it is possible to easily classify the gastric lymphomas. We suggest that an improvement of the results could be achieved by comparing the immunophenotype of the cellular lymphomatous populations, and the AgNORs pattern with patients survival.


[Psychoacoustic tests: effects of atropine].

The efferent pathways exert a control action on the function of the cochlear nucleus and hair cells. Acetylcholine is the neurotransmitter of the centrifugal system and its action can be blocked by atropine. In order to give a contribution to the knowledge of the function of the efferent bundle and of the cochlea efficiency we examined 10 young normal subjects before and after infusion of 1 mg of atropine i.v. a battery of three psychoacoustical tests (Remote Masking, Critical Ratio and Brief Tone Audiometry). After infusion of atropine we have shown an increase of 0.25 Hz hearing threshold, an increase of RC values and a decrease of RM values. It can be concluded that the pharmacological block of the olivo-cochlear bundle determines a stiffness of outer hair cells and basilar membrane; this finding means that the atropine can inhibit the facilitating activity of the efferent system on the cochlear performance.


[Polyester treatment in rats with a dorsal air pouch: chemotaxis in lavage fluid from the pouch].

Minced polyester threads introduced into peritoneal cavity of rats cause a granulomatous inflammation with evidence of macrophage stimulation. Chemotactic agents play an important role in the inflammatory reaction; they are released locally by cells involved in inflammation. In this paper the chemotactic effect of the peritoneal and subcutaneous air pouch fluids from rats bearing the polyester inflammatory process, have been studied on PMN cells "in vitro". The fluids were obtained by washing the cavity of untreated rats or rats injected with polyester, 7 days after the injection. The chemotactic response was assayed by employing modified chemotaxis Boyden chambers (Blind Well Neuro Probe) and polymorphonuclear cells from normal rats. Quantification of the migration was calculated by chemotactic index (A/B) (B = random migration, A = chemotaxis). The results demonstrate that a chemotactic activity is present in peritoneal and subcutaneous air pouch fluids following the inflammatory process. In conclusion the chronic inflammation determines the appearance of chemotactic factors for PMN cells, in the peritoneal cavity and in the air pouch, and the air pouch is a very convenient experimental system with the particular advantage that it permits easy repeated sampling of exudate during the course of an inflammatory response.


Problems of deep foveas.

Deep or convexiclivate foveas occur in some birds, including raptors, some lizards and certain deep-sea fishes. Theories on their function are reviewed. Common to raptor and deep-sea fish foveas is a radial fibre lining, dark staining so probably optically dense, adjacent to the less refractile vitreous. The foveal curvatures and size are remarkably similar in a wide taxonomic and size range of birds and fishes. Ray plotting through traced foveal outlines suggests that sharp images will be formed beneath the centre and shoulders, with the centre image enlarged enough to account for the high acuity of raptors. Deep foveas will also exaggerate eccentricity of off-centre images of a point source, such as deep-sea fishes may meet. Despite similarities in foveal shape, the receptors differ widely between species. Raptors, and notosudid fishes, have short cones. Searsid fishes have long single rods. Howella and Bajacalifornia have multiple bank rods, more in the fovea than the periphery. Those of Howella are shown to be multiple inner-outer segment complexes rather than interrupted single rods. Implications of foveal and receptor features are discussed.


Ophthalmology in the undergraduate curriculum. A review in Queensland.

Three hundred and sixty-nine postal questionnaires related to the content and teaching of the undergraduate medical course were sent to Queensland city and provincial ophthalmologists, general practitioners, physicians and surgeons. The return rate was 53%. The views of the four groups were similar in most respects. The majority of respondents thought that ophthalmology should be taught as a separate course. They stated visits to operating theatre and eye casualty unit should be included in the course. Ophthalmologists and general practitioners stated the course should be longer, physicians thought it should remain the same length and surgeons felt less time should be dedicated to the teaching of ophthalmology. Topic areas regarded as essential were the acute care areas: trauma; glaucoma; infection; use of an ophthalmoscope; acute visual loss; and red eye. Squint was also regarded as an essential area by ophthalmologists.


The effect of topical corticosteroids on laser-induced peripheral anterior synechiae.

To assess the influence of different topical steroid agents and a non-corticosteroid medication after Argon laser trabeculoplasty (ALT) on the development of peripheral anterior synechiae (PAS) and the reduction of intraocular pressure (IOP). In two separate prospective, randomised, group-controlled studies, topical fluorometholone 0.1% (FML Liquifilm) was compared with dexamethasone 0.1% (Maxidex) (Study A), or naphazoline hydrochloride 0.1% (Albalon) (Study B) after ALT for chronic open-angle glaucoma, with particular reference to the formation of PAS and the IOP response. In Study A (N = 109) eyes treated with Maxidex had a significantly higher incidence of PAS than those treated with FML - 45% compared with 22%, P < 0.05 (normal deviate test). In Study B (N = 75) the incidence of PAS was equal in eyes treated with FML or Albalon (23%). In the two studies combined (N = 184), the development of PAS was associated with a significantly lower mean response of IOP to ALT - 1.47 mmHg compared with 3.22 mmHg, 0.01 < P < 0.05 (Student t-test). The incidence of PAS after ALT is significantly higher with the post-laser use of Maxidex than with FML, and is the same for Albalon as for FML. The therapeutic benefit of ALT is significantly reduced if PAS develop.


A retinitis pigmentosa register for western Australia.

This paper describes the main elements of the Western Australian retinitis pigmentosa register including details of the data stored on the register, aspects of the coding systems used and some description of the tests employed in diagnosis of retinitis pigmentosa. The register is family based and contains data on affected individuals and on their unaffected relatives. As at November 1991, the register contained data for 391 individuals from 207 separate families. Of the 391 individuals, 240 had definite or probable retinitis pigmentosa and 26 were possibly affected. The remainder were unaffected family members. In many cases, both affected and unaffected family members are being studied serially and the register is designed to store and easily retrieve serial data to allow study of disease progression for individuals and within families.


Optic neuritis in children with poor recovery of vision.

We reviewed the records of 10 children with optic neuritis in whom recovery of vision was poor or incomplete. Our cases were otherwise similar to those described in previous studies in that they were always bilateral, often accompanied by a viral prodrome (seven of 10), and usually associated with disc oedema (seven of 10). Seven of twenty eyes had a final visual acuity of 6/60 or worse and only one patient regained 6/6 vision in either eye. In three patients the best vision in either eye was 6/60 or worse. Recovery of vision was often slow, taking up to six years. Five of 10 patients have developed multiple sclerosis (MS), and one child had acute disseminated encephalomyelitis (ADEM) with optic neuritis. Optic neuritis in children does not always carry a good prognosis for recovery of vision; however, the failure of vision recovery in a short period of time does not necessarily indicate a poor outcome. Some children with optic neuritis develop MS, which can develop even when optic neuritis follows a viral illness.


Haemangiopericytoma of the orbit.

Orbital haemangiopericytomas are ideally managed by complete surgical excision in the first instance. This is frequently not achieved, because difficulty in making the diagnosis preoperatively results in incisional biopsy and the highly vascular nature of the tumour makes complete excision difficult. A series of 12 patients with orbital haemangiopericytoma seen over a 23-year period is presented. The following combination of clinical and radiological features is suggestive of haemangiopericytoma. 1. Painless non-axial proptosis with downward displacement of the globe. 2. Intermittent upper lid swelling. 3. A soft, superiorly located mass with poorly defined borders, especially with a blue hue. 4. A superiorly located, rounded or elongated extraconal mass on CT, isodense with brain, with smooth, well-defined borders and moderate to marked enhancement with the injection of intravenous contrast medium. 5. Significant blush in all three phases of carotid angiography, without prominent arteriovenous shunting. Once haemangiopericytoma is suspected, complete surgical excision is recommended.


Paediatric dacryocystorhinostomy.

The results of 95 dacryocystorhinostomy procedures in 78 paediatric patients are reviewed. The success rate was 77%. Factors influencing success and failure are discussed.


Superior sagittal sinus thrombosis--an unexpected cause of papilloedema.

Two patients with superior sagittal sinus thrombosis are reported. Papilloedema was an important diagnostic sign in both cases. Magnetic resonance imaging (MRI) provided the diagnosis in each case and proved to be the investigation of choice. Superior sagittal sinus thrombosis should be considered in the differential diagnosis of papilloedema as it is both potentially fatal and probably underdiagnosed.


Preservation of residual field after surgical lowering of intraocular pressure.

A 52-year-old Caucasian woman with pigmentary glaucoma underwent a left cyclodiathermy procedure for raised intraocular pressure (IOP) in 1962 and right sclerectomy the following year. Over 25 years later a low IOP was still maintained though pilocarpine was needed in the left eye. After a left cataract extraction with insertion of a posterior chamber lens, field loss had not progressed very much in either eye though the visual acuity was reduced in the more affected eye. Both drainage operation and a cyclodestructive procedure controlled IOP over a long period of time in this patient and were associated with only a very gradual progression of field loss.


[Preparation and utilization of BrdU antiserum. II. Detections of BrdU labelling chromosome and DNA using BrdU antibody technique].

Utilizing highly specific BrdU antiserum, via the second antibody conjugated with enzyme and DAB-H2O2 or AEC-H2O2 colorific methods, the BrdU marked regions of CHO and human chromosomes were detected clearly. The BrdU-DNA prints (pg) on nitrocellulose membranes were also showed with similar methods.


[Studies of distribution of subtypes of transferrin, group-specific component, alpha 1-antitrypsin in gastric cancer patients from Shanghai area].

The genetic polymorphism of transferrin (Tf) and alpha 1-Antitrypsin (alpha 1-AT) in 202 gastric cancer patients and 202 controls in Shanghai Hans were analyzed by isoelectric focusing. In transferrin, the phenotype frequency of C1C1 and gene frequencies of c1 and c2 were 0.3713, 0.5718 and 0.4109 respectively in gastric cancer patients, and 0.5149, 0.6782 and 0.2970 separately in controls. The frequencies of the c1c1 phenotype and the C1 gene increased significantly (p < 0.01) among the controls, while the frequency of the C2 gene greatly increased (p < 0.01) among the patients. The frequencies of the C2C2 in gastric cancer patients and controls were 0.2228 and 0.1436 respectively, showing a significant difference between them (p < 0.05). The genetic polymorphisms of group-specific component (Gc) in 200 gastric cancer patients and 200 controls in Shanghai Hans were studied with isoelectric focusing followed by immunofixation. The Gc1F1F phenotype frequency and 1Fgene frequency were 0.22 and 0.4375 in gastric cancer patients, and 0.14 and 0.3600 in controls, respectively. The frequencies of the Gc1F1F phenotype and the 1F gene were significantly increased (p < 0.05) among the patients. In alpha 1-Antitrypsin, there was no significant difference in phenotype frequencies and gene frequencies between the patients and controls. The data reported here indicate that the gastric cancer is associated positively with TfC2 and Gc1F, and negatively with TfC1. These facts support the notion that cancerogenesis is a multi-step process controlled by multifactorial inheritance.


[Synaptonemal complexes of human spermatocytes and foetal oocytes observed by electron microscopy].

The electron microscopical studies on the synaptonemal complexes (SC) of Chinese spermatocytes and foetal oocytes have been made and used to construct SC karyotype with a modified surface micro-spreading technique. We also confirm the SC formation between the short arms of the X chromosome and Y chromosome as well as secondary association between the distal tips of Xq and Yq at pachytene. Moreover, attentions are paid to the morphology of the XY pairs and five pairing type of XY pairs. In our preparations, a rare triploid spermatocyte was observed and triple pairing was described. Some possible significances of the triple pairing of triploid spermatocyte in relation to the human chromosomal abnormalities have been supposed.


[Isolation and characterization of prolactin messenger RNA from bovine anterior pituitary glands].

The messenger RNA was extracted from bovine anterior pituitary glands and was purified by oligo(dT)-cellulose chromatography. The length of the mRNA was 1,200 nucleotides measured by agarose gel electrophoresis containing methylmercury hydroxide. The bovine prolactin (PRL) mRNA was confirmed by Northern blot analysis of the mRNA with gamma-32P labelled synthetic oligonucleotide probes based on the partial amino acid sequences of bovine PRL and could stimulate the incorporation of 35S-methionine into protein in the rabbit reticulocyte cell-free system. The translation product of bovine PRL mRNA was immunoprecipitable by rabbit anti-ovine PRL anti-antibodies. The molecular weight of the translation product corresponding to the bovine PRL precursor was estimated to be approximately 25,000 by SDS polyacrylamide gel electrophoresis and autoradiograph.


[Iterative estimation of variance components for a mixed model with a numerator relationship matrix: apply to compute K of variance ratio in BLUP and genetic parameters].

An iterative procedure is presented for the estimation of variance components in the mixed model with a numerator relationship matrix from non-balanced data. This method has been shown to be feasible if only designing matrix for fixed effects of the mixed model and Henderson method 3 hypothetic condition of the fixed effect to compute correctly the reduction in sum of squares, we would obtain more precise estimation of variance components, furthermore, the sizes of the initial value K could not affect to last result of the iterative solution. This is because in the iterative process may adjust automatically by designing matrix. Also, the variance components could apply not only to compute variance ratio in BLUP, but also to estimate genetic parameters.


[Nucleotide frequency analysis of bacteriophage phi X174 genome].

Detailed analysis was carried out on the nucleotide frequencies in the genome of bacteriophage phi X174. In coding regions the distribution of four nucleotides at the three codon sites was highly non-random, and the patterns were conspicuously similar among the genes, implying that it might be a feature of the genome. An index c, which was based on the chi 2 statistics of the site by base 3 x 4 table, was introduced to reveal he extra information embedded in the coding region, and also to discriminate between coding and non-coding open reading frames. Doublet deviation analysis showed that deviation patterns were not significantly related to the codon sites in coding regions, and were also very similar in different genes. Possible mechanisms of such similarity are discussed.


The antimicrobial activity of non-antibiotics: interactions with antibiotics.

The following are considered: the direct antibacterial activity of non-antibiotics: resistance-reversing agents and resistance-inducing agents; the antibacterial and antiparasitic activity of non-antibiotics: psychotropics, antihistaminics, antineoplastics, allopurinol, hyperbaric oxygen, etc.; as well as the antibacterial activity of agents modifying the urinary pH; the influence of non-antibiotic drugs on the pharmacokinetic parameters of antibiotics; the anti infectious activity of non-antibiotics through their effects on host defences, immunity and inflammation, and finally the use of non-antibiotics to decrease the toxicity of anti-infectious agents.


Antiplasmid activity: loss of bacterial resistance to antibiotics.

The antiplasmid activity of tricyclic compounds, e.g. phenothiazines, dibenzoazepines, dibenzocykloheptene derivatives and some stereoisomers, was shown on E. coli in vitro. Some ring-substituted phenothiazine and cannabis derivatives had only an antibacterial effect. Promethazine, a selected phenothiazine, cured antibiotic resistance and lactose fermentation of E.coli, tumour inducing ability of Agrobacterium tumefaciens and nodule formation of Rhizobium meliloti. Plasmids of different E.coli strains were eliminated with varying frequency. The antiplasmid activity of the compounds can be due to the increased membrane permeability. Inhibition of DNA gyrase and complex formation with the supercoiled form of plasmid DNA can lead to the cessation of plasmid replication in the bacterial cells. In addition, in vivo plasmid curing was demonstrated at a low frequency.


The modulation of the bacteriolytic effect of beta-lactam antibiotics by non-antibiotics.

The addition of cationic proteins such as lysozyme, ribonuclease and cytochrome C enhanced the beta-lactam-induced bacteriolysis of staphylococci measured as release of wall label or by optical density. The treatment of staphylococci with penicillin plus cytochrome C resulted in a reduced viability of bacteria compared with those treated with penicillin alone. The wall autolysis and the penicillin-induced bacteriolysis of staphylococci were enhanced by the lysosomal enzyme cathepsin C. The penicillin-induced bacteriolysis was also enhanced by the D-amino acids D-alanine and D-methionine, while the comparable L-amino acids did not reveal any activity. On the other hand, some polyanionic substances were able to suppress the penicillin-induced bacteriolysis. Radiochemical and electron microscopic studies revealed the participation of bacterial wall autolysins in the first steps of degradation processes of staphylococcal walls within murine bone marrow-derived macrophages.


In vitro antimicrobial activity of 18 phenothiazine derivatives: structure-activity relationship.

The antimicrobial activity of 18 phenothiazine derivatives has been evaluated against aerobic and anaerobic bacteria. The mean of the MIC reveals five active drugs against the bacterial strains tested: thioridazine, chlorpromazine, trifluoperazine, fluphenazine and triflupromazine. The other derivatives are less active and some show no antimicrobial activity. It seems that in vitro antibacterial activity alone is insufficient to explain certain intestinal diseases linked to the use of these molecules in psychiatry.


The antimicrobial activity of non-antibiotics. Report from a congress on the antimicrobial effect of drugs other than antibiotics on bacteria, viruses, protozoa, and other organisms.

In May 1990 the 1st International Conference on Antimicrobial Activity of Non-Antibiotics was held in Copenhagen, Denmark. More than 200 scientists were drawn from 34 countries and from all five continents. Papers were presented dealing with the antimicrobial activity of different synthetic and natural compounds, and interactions between antibiotics and non-antibiotics. The conference pointed to the unusual properties of non-antibiotic drugs. These effects might create unexpected therapeutic possibilities and lead to new basic insights.


Microcirculation of the rat pancreas, with special reference to the insulo-acinar portal and insulo-venous drainage systems: a further scanning electron microscope study of corrosion casts.

Microcirculation in rat pancreas was studied by scanning electron microscopy of resin vascular casts and by light microscopy of India ink-injected and sectioned tissue samples. In the scanning survey of the casts, islets larger than 30 microns in diameter counted no less than 400 in the adult rat pancreas. They were located either in the exocrine lobules (intralobular islets, counting 210 or more), or in the interlobular tissue spaces and along the secretory ducts (extralobular islets, 190 or more). Every islet received arterial blood via one or more proper arterioles. These afferent vessels first divided in the peripheral zone of the islet consisting of A and D cells and later extended deeper to form a capillary network in the islet core consisting of B cells. Blood originating from this capillary network left the islet via efferent vessels. This microvascular pattern may favor the regulation by A and D cells over the secretory activity of B cells. The intralobular islets gave their efferent vessels to the capillary bed of the exocrine tissue of the lobule, thus forming an insulo-acinar portal system. When the lobule was relatively small in size, superfluent blood was led directly to veins via insulo-venous drainage vessels. The intralobular islets sometimes issued one or more long translobular portal vessels reaching adjacent lobules. The extralobular islets issued either insulovenous vessels draining to interlobular veins, or extralobular insulo-acinar portal vessels supplying adjacent lobules.


Two components of the pineal organ in the mink (Mustela vison): their structural similarity to submammalian pineal complexes and calcification.

The pineal complex in the mink (Mustela vison) consists of a larger ventral and a smaller dorsal pineal. Both organs contain pinealocytes, neurons, glial cells, nerve fibers and synapses in an organization characteristic of nervous tissue. The cellular elements are arranged circularly around strait lumina. These lumina correspond to the photoreceptor spaces of submammalian pineals. A 9 + 0-type cilium marks the receptory pole of the pinealocytes which may form an inner-segment-like dendrite terminal in the pineal lumina. The cilia correspond to outer segments which form photoreceptor membrane multiplications in the pineal of submammalians and in certain insectivorous and mustelid mammals (bat, hedgehog, ferret). Axonal processes of the pinealocytes contain synaptic ribbons and terminate on intrapineal neurons of both organs. This pattern represents a neural efferentation of the pineal nervous tissue. The axonal processes of pinealocytes also form neurohormonal endings which pierce the perivascular limiting glial membrane in the ventral as well as in the dorsal pineal. The upper pineal ("epipineal") of the mink may correspond to the parapineal, frontal, or parietal organs of submammalian pineal complexes. Both pineals are encapsulated by the meningeal tissue of the brain stem. Afferent vasomotor axons of the meninges innervate smooth muscle cells of pineal arterioles. There are corpora arenacea in the pineal arachnoid and in the pineal nervous tissue, primarily in the ventral pineal. The localization of calcium ions detected around the membrane of pineal cells by pyroantimonate cytochemistry suggests membrane activity as the source of the calcium ions. The accumulation of calcium by the pinealocytes may be due to their neurosensory character. The mink is the first animal described to have both intrapineal and meningeal concrements like the human pineal.


Semiquantitative analysis of the effects of fixation and paraffin embedding on immunoreactivity of renal basement membranes to a monoclonal antibody against type IV collagen.

The effects of formalin fixation and paraffin embedding on the immunoreactivity of human kidney to a monoclonal anti-type IV collagen antibody (JK-199) were examined semiquantitatively by a modified enzyme-linked immunosorbent assay (ELISA). The intensity of immunoreactivity in paraffin sections of the tissue fixed overnight with 10% formalin was approximately 70% of that in frozen sections. Immunoreactivity reduced to this extent did not impair the specific staining of basement membranes. Paraffin sections of tissues fixed 2 days showed 50-60% of the reactivity in the frozen sections of the tissue fixed overnight; the basement membranes in Bowman's capsules were stained positively, but those in other sites were not. The paraffin sections of tissues fixed 7 or 14 days showed no specific immunostaining. The immunoreactivity for type IV collagen in the basement membranes was restored after treatment with pronase E. The immunoreactivity after the enzymatic treatment was about 150% of that in the frozen sections of the overnight fixed specimens.


Localization of hyaluronic acid, chondroitin sulfate, and CD44 in rabbit cornea.

To demonstrate the localization of hyaluronic acid (HA) in rabbit cornea, the biotinylated HA-binding region, which specifically binds to the HA molecule, was applied to the tissue. Localization of chondroitin sulfate (CS) and CD44, a possible cell surface receptor for HA, were also examined by immunohistochemistry. The stainability of HA changed depending on the fixatives used. Reaction products for HA were distinctly detected in epithelial cells and stromal keratocytes, but faintly in the extracellular matrix of the stroma when unfixed cryosections were applied. No positive reaction was found in the endothelium, except that the positive deposits formed a continuous layer on the apical surface of the endothelium. Electron microscopy using samples fixed with 2% paraformaldehyde revealed gold particles indicating HA labeling the intercellular space of the epithelium and stromal extracellular matrix. No intracellular deposition was detected in epithelial cells, whereas the gold labeling was seen in vacuolar structures of stromal keratocytes. Immunodeposits for CS were intensely localized in the epithelium and stroma, and weakly in the endothelium. Immunoreactivity for CD44 was found in the epithelial, endothelial and stromal cells. In particular, immuno-deposits for CD44 were detected in basal parts of epithelial cells, while they were localized in the apical surface of endothelial cells. These results suggest that HA is synthesized in and secreted from epithelial and stromal cells of rabbit cornea, while the localization of HA in the apical surface of the endothelium is closely associated with that of CD44. Moreover, the presence of CS in corneal tissue may play a role in its transparency, as has been suggested for keratan sulfate and dermatan sulfate.


Structure and distribution of the lymphatic vessels in the parietal pleura of the monkey as studied by enzyme-histochemistry and by light and electron microscopy.

The entire distribution of lymphatics in whole mount preparations of the Japanese monkey was studied using the enzyme-histochemical technique reported by KATO et al. (1990, 1991). In this staining, the lymphatic endothelium was colored dark brown by its positive 5'-nucleotidase activity, while most blood vessels (especially arterioles) were colored blue due to their positive alkaline phosphatase reaction. The whole mount preparations of the pleura treated enzyme-histochemically clearly indicated the distribution, branching patterns and running courses of lymphatic vessels. They revealed numerous short blind-ending knobs which represented the initial portions of lymphatics. These knobs were seen near the surface of the parietal pleura along its entire extent. In the costal and diaphragmatic pleura, the lymphatics ran parallel to the intercostal muscle fibers, but perpendicular to the tendinous and muscular fibers of the diaphragm; they formed ladders, independent of the courses of blood vessels. In the mediastinal pleura, lymphatic vessels showed a tree-like branching accompanying blood vessels. Under the light microscope, toluidine-blue stained semithin sections revealed the initial part of lymphatics as a small irregularly outlined cavity (7-10 microns in diameter) surrounded by a dense connective tissue. This lymphatic dilation was sometimes located close to a thin mesothelial layer. Such a structure suggesting a "stoma" was seen near the attachment of the muscular diaphragm to the sternum and along the borders of the ribs. Transmission electron microscopy revealed an occasional interruption in the mesothelium. This stoma continued to a submesothelial cavity whose base comprised an attenuated endothelium of an extended lymphatic vessel.


Three-dimensional changes in direction and interrelationships among enamel prisms in the dog tooth.

This study describes the three-dimensional features of enamel prisms and their arrangement in dog teeth. Tangential semithin sections of demineralized tooth germ were serially cut from the enamel surface to the enamel-dentin junction. Straight rows of enamel prisms parallel or perpendicular to the meridian were selected at the enamel-dentin junction; these prisms were reconstructed from micrographs with a personal computer. Near the enamel-dentin junction, the arrangement of enamel prisms appeared regular. Viewed from the enamel surface, the cut-ends of the enamel prisms that were parallel to the meridian at the enamel-dentin junction appeared as a sine curve, with 16 enamel prisms forming one period. The enamel prisms in a row perpendicular to the meridian were parallel to each other and deflected to the left or right from the enamel-dentin junction. Away from the enamel-dentin junction, the periodicity of the prisms gradually disappeared. The sine curve formed by the cut-ends of prisms in a row parallel to the meridian became irregular, and prisms in rows perpendicular to the meridian crossed each other. The semithin sections showed belt-like zones arranged perpendicular to the meridian. Each belt-like zone consisted of enamel prisms oriented in the same direction, those in neighboring zones being oriented in opposite directions. The disappearance of the regular arrangement of prisms was related to changes in their location in the belt-like zones.


Electron microscope observations on the formation of primitive villi in rat small intestine with special reference to intercellular junctions.

Formation and fusion of intraepithelial cavities have long been considered an essential process in the histogenesis of the intestinal mucosa. By electron microscope observation of thin sections and freeze-fracture replicas of the small intestine of rat fetuses, we first demonstrated the initial steps in the formation of intraepithelial cavities: A focal tight junction (macula occludens) was formed in the abluminal part of the epithelium, after which the membrane of an intracellular cavity was fused with that of the focal tight junction to form an intercellular (intraepithelial) cavity enclosed by a zonula occludens. The present study also revealed that gap junctions appeared and enlarged simultaneously with the formation of primitive villi and differentiation of absorptive cells. These gap junctions gradually came to be confined in the epithelium of intervillous regions where proliferation and differentiation of epithelial cells took place. Absorptive cells in villi rarely had gap junctions. These results suggest that tight and gap junctions play important roles in the histogenesis of the intestinal mucosa, and in the proliferation and differentiation of epithelial cells.


Immunoelectron microscopic observation of calcitonin gene-related peptide (CGRP)-positive nerves in the dental pulp of rat molars.

The ultrastructure of nerves containing immunoreactivity for calcitonin gene-related peptide (CGRP) was investigated in the dental pulp of rat molars. The immunoreactivity was recognized predominantly in unmyelinated nerve fibers, and sparsely in a few myelinated fibers. It was localized throughout the axoplasm, as well as in the large cored vesicles. Small clear vesicles and mitochondria were free of the immunoreaction. The CGRP-immunoreactive nerves were frequently observed to terminate, being devoid of Schwann cell investment, in the vicinity of blood vessels in the coronal pulp, suggesting that CGRP may be involved in the regulation of pulpal blood flow. Moreover, CGRP-immunoreactive axon terminals containing numerous small clear vesicles, a few large cored vesicles and mitochondria were recognized in contact with the cell bodies of odontoblasts and their processes in the dentinal tubules. Although specialized synaptic ultrastructures were not recognizable, a functional association of CGRP nerves and odontoblasts was suggested. Thus, CGRP in the dental pulp appears to have multiple functions, including vascular regulation and sensory transduction.


A comparative study of myogenic cell invasion of the avian wing and leg bud.

The borders of myogenic cell invasion of avian wing and leg buds were determined using the interspecific grafting technique between quail and chick embryos. Distal parts of quail limb buds were grafted ectopically into the coelomic cavity of chick embryos. The presence or absence of skeletal muscle was investigated in histological sections of the reincubated grafts. A comparison between the borders of myogenic cell invasion of the wing and leg buds showed that the differences in the position of the distal most muscles in the adult avian limbs could be a consequence of the cranio-caudal sequence of development.


A two-step alignment method for 3D computer-aided reconstruction based on fiducial markers and applied to mouse embryonic hearts.

An accurate three dimensional computer reconstruction of microscopic biological objects or distribution of molecules identified on serial sections must solve two major problems: 1) the alignment of sections using adequate extrinsic references (fiducial markers); 2) the impossibility of observing these references and the cellular or molecular structures in the microscope at the same magnification. To provide extrinsic references for objects embedded in soft media, we have modified and simplified the charcoal-paraffin method described by Langemeijer and Simons (1973). It consists of drilling three or four small holes into the paraffin block, sealing this block at the extremity of a glass holder and, from the other extremity of the holder attached to a rubber hose, aspirating a liquefied mixture of charcoal-paraffin to fill these cylindrical holes. An alignment procedure was developed using serial sections of mouse embryonic hearts with bromodeoxyuridine-labelled DNA synthesizing cells. From each fourth section, two sets of contours have been drawn and digitized: 1) at low magnification (about 40x), embryo body wall, heart, neural tube and extrinsic reference marks (black dots); 2) at higher magnification (240-300x): heart contours alone (without extrinsic references, but with individual labelled cells). Different operations of the computer-aided alignment, as well as checking of results by inverse alignment, are described in detail. This two-step alignment method offers a practical, efficient compromise between: a) purely subjective alignment based only on tissular landmarks interpreted by the operator; b) ideal perfect alignment based not only on adequate references, but on computerized correction of section deformation, as well.


Origin of primordial germ cells, as characterized by the presence of nuage, in embryos of the teleost fish Barbus conchonius.

The presence, location and morphology of cells containing nuage, an ultrastructural characteristic of primordial germ cells (PGCs), is described from the moment of first morphological recognition of PGC (around 100% epiboly) in embryos of the teleost fish Barbus conchonius. Thus characterized cells were studied in relation to their cellular contacts with somatic germ layer cells, possibly involved in the determination of PGCs. The results show that from the very moment that cells, likely to be PGCs, can be light microscopically identified with morphological and positional criteria (from 10 h post fertilization (p.f.) onwards), they contain nuage near the nuclear envelope, which is a strong indication of their PGC-identity. During the studied period (9-12 h and 24 h p.f.) nuage-containing cells seem to translocate from the mesoderm towards the yolk syncytial layer (YSL). These PGCs usually appear not to be directly connected with the YSL but to remain separated from the YSL by one or more endodermal extensions, at least up to 12 h p.f. Also at 24 h p.f. somatic cells separate the PGCs from the YSL.


Morphological indications of secretion by uterine epithelial cells and changes in the ratio between acidic and basic uterine proteins during early pregnancy in Chinese Meishan pig.

In Chinese Meishan pig embryonic mortality appears relatively low compared to European breeds. Most of embryonic loss in pig is believed to take place during early pregnancy. It is of interest to know possible specific features associated with low mortality. Therefore, the ultrastructure of the endometrial epithelium of Meishan pig was studied between Days 4 and 12 of pregnancy, and compared with earlier results on Yorkshire/Dutch Landrace interbreed (Y/DL). Furthermore, total protein and the relative amounts of acidic and basic proteins were determined in the uterine flushings, and compared with earlier results on Dutch Landrace (DL). As holds for European breeds, uterine glandular and luminal epithelium have to be considered as functionally different cell populations. Their morphology differs and suggests the synthesis of different secretory products. The periods of secretion are not the same: the luminal epithelium shows signs of product release during the whole period studied, the glands deliver their secretions from Day 12. This is correlated with a sudden increase in total protein in uterine flushings. Between Days 4 and 12, the relative amount of acidic proteins decreases from 92% to 47% in DL and from 88% to 38% in Meishan, resulting in a shift from acidic protein dominance to a mild dominance of basic proteins in both breeds, but most prominent in Meishan.


Geometrical differences between neck motoneurons located in the brainstem and spinal cord in the mallard Anas platyrhynchos L.; a Golgi study.

The dendrite geometry of neck motoneurons located in the upper cervical cord and lower brainstem of the mallard was studied in Golgi silver impregnated material. Measurements were obtained from camera lucida drawings and concerned the extent and orientation of dendritic trees. Dendrites were found to be oriented predominantly parallel to the sagittal plane, and projections were asymmetric in the dorso-ventral direction. Comparison between motoneurons located in the supraspinal nucleus of the brainstem and motoneurons in the ventral horn of the upper spinal cord showed that dendrites of motoneurons in the first cell group branch more often than those of neurons in the latter group. In addition, dendritic trees of ventral horn motoneurons preferentially project into the field dorsal to the cell body.


The developmental mutants of Xenopus.

The 32 developmental mutants found in our laboratory have appeared in the course of the genetic analysis of adult Xenopus laevis issued from nuclear transfers, and of their progeny. These experiments originally were devised more than thirty years ago to test whether somatic nuclei had undergone irreversible changes during differentiation or whether they had remained totipotent. In the majority of cases, the mutations were carried by wild-caught imported frogs or their laboratory-bred progeny; however, the precise origin of several mutations has not been determined. The mutants have been subdivided into three classes: Maternal effect mutants. Three such mutants have been found; they affect the development of all the embryos from homozygous mothers independently of the genotype of the father. Developmental lethals. This class comprises 19 mutants, among which are included the previously called "autonomous lethals" caused by general metabolic defects that affect all parts of the embryos as well as the tissue- and organ-specific lethals, in several of which death occurs as a direct consequence of the anomalies. Developmental non-lethals. These are 10 mutants, suffering from specific defects not essential for survival. In addition, nucleolar mutants as well as mutants found in different species of Xenopus and mutants of Xenopus laevis found in other laboratories are also mentioned. The last part consists of an alphabetical description of the mutant phenotypes including more detailed analyses which have been carried out on several of them.


Effects of dentin proteins, transforming growth factor beta 1 (TGF beta 1) and bone morphogenetic protein 2 (BMP2) on the differentiation of odontoblast in vitro.

We have studied the effects of dentin proteins, of Transforming Growth Factor beta 1 (TGF beta 1) and Bone Morphogenetic Protein (BMP2) on the differentiation of odontoblasts in vitro. The total EDTA-soluble fraction of dentin proteins, prepared from rabbit incisors was further separated by chromatography on DEAE-Cellulose and heparin-agarose columns. While the total EDTA-soluble fraction of dentin had no effect on cultured dental papillae, fractions retained on both columns were able to initiate functional differentiation of preodontoblasts of isolated day-17 first lower mouse molar dental papillae cultured in vitro. TGF beta 1 and BMP2, both stimulated the matrix secretion by dental papillae cells. TGF beta 1 and BMP2, combined with the inactive total EDTA-soluble fraction, stimulated odontoblast differentiation. An active fraction retained on DEAE-Cellulose completely lost the inductive activity after incubation with a neutralizing anti-TGF beta antibody. These results demonstrate that a TGF beta-like molecule present in dentin could interact with some component which acts as a modulator of its activity on the initiation of the cytological and functional differentiation of odontoblasts.


Endogenous epidermal growth factor regulates the timing and pattern of embryonic mouse molar tooth morphogenesis.

The tooth organ provides a model for discrete patterns of morphogenesis over short periods of developmental time. Studies were designed to test the hypothesis that endogenous epidermal growth factor (EGF) functions to regulate multiple cusp molar tooth morphogenesis during embryonic mouse development. The relative levels of endogenous EGF and EGF receptor (EGFR) transcripts were determined in both enamel organ epithelia and dental ectomesenchyme by reverse transcription-polymerase chain reaction (RT-PCR) assays. EGF and EGFR were localized by immunohistochemistry; both antigenic determinants were demonstrated on the same odontogenic cells in cultured tooth explants. To examine EGF-mediated signal transduction, cap stage mouse molar tooth organs (E16) were cultured in serumless, chemically-defined medium as either (i) controls, or supplemented with (ii) tryphostin (an EGF receptor kinase inhibitor), (iii) tyrphostin plus exogenous EGF, and (iv) exogenous EGF. Antisense oligodeoxynucleotide (ODN) strategy was used to investigate the functions of endogenous EGF employing (i) non-treated control, (ii) sense ODN control, (iii) antisense ODN, (iv) exogenous EGF, (v) sense ODN with exogenous EGF, and (vi) antisense ODN with exogenous EGF. Tyrphostin inhibited DNA synthesis and produced a significant decrease in the volume of the explants. These effects were recovered by addition of exogenous EGF. Antisense ODN inhibition resulted in abnormal cusp formations, decreased DNA synthesis, total DNA, RNA and protein content, and decreased stellate reticulum and tooth explant volumes. The decreased tooth size was not uniform, the most pronounced effect was in the stellate reticulum. This pattern of changes was not seen when antisense ODN treatment was supplemented with exogenous EGF. These results suggest that during cap stage of odontogenesis endogenous EGF acts to stimulate DNA synthesis, which increases the cell number of specific phenotypes within the enamel organ epithelia, and thereby regulates molar tooth morphogenesis.