Importance of tyrosine phosphatases in the effects of cell-cell contact and microenvironments on EGF-stimulated tyrosine phosphorylation.

We have compared the EGF responses of A431 cells when grown as monolayers at a variety of cell densities or as multicellular spheroids in order to investigate the effects of cell contact and 3-dimensional structure on signal transduction. Proliferation of the A431 squamous carcinoma cell line grown in our laboratory was unaffected by EGF when grown in monolayer culture. As 3-dimensional, multicellular spheroids, however, growth was stimulated by EGF. The maximum volume attainable in the presence of EGF was more than 30 times that in its absence. EGF-dependent tyrosine phosphorylation was compared under these conditions by immunohistochemistry and Western blotting. In initial experiments using published procedures, tyrosine phosphorylation was density-dependent in monolayers and undetectable in spheroids. However, the density-dependence was abolished by the addition of high concentrations of protein tyrosine phosphatase inhibitors (1 mM Zn++ and VO4(3)-). The density dependence of EGF-stimulated tyrosine phosphorylation in monolayers was, therefore, largely the result of changes in phosphatase activity rather than kinase. Using high concentrations of phosphatase inhibitors, phosphotyrosine was clearly visible by immunohistochemistry in the outermost cells of spheroids, but it was still not visible in the spheroid center. The lack of response within the spheroid was not related to the presence of EGF receptor nor diffusion of EGF. In companion experiments, we showed that staining for EGF receptor was present homogeneously throughout the spheroid and that EGF penetrated to its center under the conditions of the experiment. Thus, although an increase in tyrosine phosphatase activity was a major factor affecting tyrosine phosphorylation in the outer cells, other factors were important in the inner cells. We concluded that an increase of tyrosine phosphatase activity was the most important component of the adaptation of the EGF signal transduction system to high cell density in monolayer cultures. In spheroids, tyrosine phosphatases are also enhanced, but other factors, such as autocrine synthesis of TGF-alpha and possibly the cellular distribution of EGF receptors and cell shape, play a role.


Regulation of programmed death in erythroid progenitor cells by erythropoietin: effects of calcium and of protein and RNA syntheses.

Erythropoietin (EPO) retards DNA breakdown characteristic of programmed cell death (apoptosis) and promotes survival in erythroid progenitor cells. The mechanism by which EPO inhibits programmed death is unknown. In the well-characterized model of glucocorticoid-treated thymocytes, activation of a Ca2+/Mg(2+)-sensitive endonuclease and new protein and RNA syntheses have been found necessary for apoptosis. We examined the effects of EPO on the free intracellular calcium ion concentration ([Ca2+]i), and the roles of Ca2+ and RNA and protein syntheses on DNA cleavage in erythroid progenitor cells. The murine model of erythroid differentiation using Friend leukemia virus-infected proerythroblasts (FVA cells) was used. EPO did not affect the [Ca2+]i in FVA cells. Decreasing [Ca2+]i by extracellular Ca2+ chelation with EGTA facilitated DNA breakdown. Increasing [Ca2+]i with the calcium ionophore 4-bromo-A23187 increased DNA cleavage; however, DNA fragments generated by high [Ca2+]i were much larger than those seen in the absence of EPO or presence of EGTA. Increased [Ca2+]i also inhibited DNA breakdown to small oligonucleosomal fragments characteristic of cells cultured without EPO. However, no concentration of ionophore protected the high molecular weight DNA as did EPO. Cycloheximide inhibited DNA breakdown in a dose dependent manner in cultures lacking EPO, but two other protein synthesis inhibitors, pactamycin and puromycin, did not prevent DNA breakdown. Inhibition of RNA synthesis with actinomycin D did not prevent DNA breakdown. Cells with morphological characteristics similar to those reported in other cells undergoing programmed death accumulated in EPO-derived cultures. These studies demonstrate that although DNA cleavage and morphological changes are common to apoptotic cells, the roles for Ca2+ and protein and RNA syntheses are not universal and suggest that apoptosis can be regulated by different biochemical mechanisms in different cell types.


[Biochemical indices of liver function in workers from repair brigades in chemical plants "Organika-Azot" in Jaworz].

In order to assess the functional state of the liver in 45 repair service workers of a chemical plant producing pesticides the serum concentration and electrophoretic pattern of proteins, the concentration of bilirubin and the activity of alkaline phosphatase, gamma-glutamyl- transpeptidase, alanine and aspartate aminotransferase, and lactic and malic dehydrogenase were determined. As compared to 35 healthy controls, not exposed to noxious chemicals, a significantly lower serum protein concentration with higher percentage of gamma-globulins and lower albumins and alpha 2-globulins were observed, the serum alanine and aspartate aminotransferase activities were significantly elevated. Ultrasound examination of the hepatic structure revealed liver steatosis in 11 (24.4%) workers. The results of our study point to a discrete lesion of the liver.


CSF 5-HIAA as a predictor of treatment response in trichotillomania.

Trichotillomania is characterized by chronic hair-pulling resulting in noticeable hair loss. In a preliminary study, cerebrospinal fluid (CSF) measures in 8 medication-free, female, trichotillomania patients were compared to those of matched, normal controls. There was no difference between patients and controls in measures of CSF cortisol, 5-hydroxyindoleacetic acid (5-HIAA), homovanillic acid (HVA) and 3-methoxy-4-hydroxyphenylglycol (MHPG). CSF measures did not correlate with measures of trichotillomania symptomatology. However, degree of response to treatment with serotonin re-uptake inhibitors significantly correlated with baseline CSF 5-HIAA. This suggests that central serotonin turnover is specifically relevant to treatment response to serotonin re-uptake inhibitors in trichotillomania.


[Anti-HCV antibodies against the structural and nonstructural antigens of hepatitis C in people seropositive in the second generation test].

Forty six anti-HCV repeatable positive (the 2nd generation Abbott EIA test) adults, among which 26 were blood donors and 19 were patients with chronic hepatitis, were tested using Abbott HCV EIA Supplemental Assay. The test renders possible to conduct separate investigation antibodies against structural and nonstructural antigens of HCV. In 44/45 (97.8 per cent) cases was an agreement of the results with the 2nd generation test. In the most of cases (80 per cent) both types of antibodies were present. However, in 20 per cent of blood donors the antibodies against structural HCV antigen were not present whereas all the chronic hepatitis patients had these type of antibodies.


Influence of atrial fibrillation on coagulo-fibrinolytic markers in patients with cerebral infarction.

To examine the influence of atrial fibrillation (Af) on stroke onset, we measured the plasma D-dimer level, thrombin antithrombin III complex and plasmin alpha 2 antiplasmin complex (PAP) in 46 stroke patients with Af and 87 stroke patients without Af. These marker levels were significantly higher in Af patients with stroke than in those without stroke (n = 16), and thus do not seem to be affected by Af alone. Abnormal values were also more frequent in acute Af stroke patients with visible occlusion of the major cerebral artery than in those without Af. The D-dimer and PAP levels in all Af stroke patients in the younger-aged patients (< or = 64 years) were significantly higher than those without Af, but not noted in the older-aged group (> or = 75 years). These results suggest that the D-dimer and PAP levels in younger-aged patients with Af indicate the existence of cerebral emboli due to Af.


Long-term prostacyclin--treatment acts on the DNA and RNA content of rat gastric (antral and fundic) mucosa dose-dependently.

The changes in gastric (antral and fundic) mucosal DNA and RNA contents were investigated in long-term (80 days) treatment of rats with orally administered prostacyclin. Prostacyclin caused a dose-dependent, significant increase of the DNA levels in both parts of the gastric mucosa together with a significant thickening of the fundic mucosa. The observed changes (decrease) of the RNA/DNA ratio in the fundic as well as antral mucosa, are interpreted as a sign of accelerated cell renewal, i.e. hyperplasia.


[Ophthalmic zoster in clinical material from the clinic of infectious diseases PAM in Szczecin during 1985-1989].

Clinical course, early and late complications as well as treatment of ophthalmic zoster in 70 hospitalized patients are presented. Authors pay the attention to early treatment (general and topical) with Zovirax in prevention of ocular complications.


[Concentration of prealbumin, ceruloplasmin, alpha-macroglobulin and haptoglobin in blood serum of patients with chronic non-A, non-B hepatitis treated with hemodialysis for chronic renal failure].

In 25 patients with chronic renal failure, treated with haemodialysis (13 patients with chronic non-A, non-B hepatitis, and 12 cases without evidence of hepatocellular damage), and in 20 healthy persons, blood serum concentrations were determined of prealbumin, ceruloplasmin, alpha 2-macroglobulin, and haptoglobin. It was found that the concentrations of these proteins in both subgroups of patients were not significantly different. The concentration of prealbumin was higher, and that of alpha 2-macroglobulin and haptoglobin was significantly lower in comparison with healthy subjects.


[Diagnostic value of prostatic specific antigen (PSA) in comparison to prostatic acid phosphatase (PAP) in prostatic cancer and adenoma].

The diagnostic value of the tumour markers: PSA, PAP and AcP was studied before treatment in 379 men (47 with prostatic cancer--PC, 306 with benign hyperplasia--PBH, and 26 healthy subjects--control group CG). PSA was determined by the enzymoimmune method, and the phosphatases were evaluated by the spectrophotometric method. Raised level of PSA was found in PBH--the highest value--23.3 ng/ml. After accepting the cutting off values (1.9 ng/ml and 23.3 ng/ml), even in 93% of patients with PC, the level of PSA exceeded the second of those values. A significant growing tendency was found of PSA together with the degree of clinical progression of PC (in stages C and D--in 100% of patients). PSA, as compared with the phosphatases, is a much more sensitive biochemical marker, exceeding them many times in sensitivity.


[Quality of life as a catch phrase and promoter in the pharmaceutical industry].

Quality of life (QL) is an expression of the degree to which a tumor patient is satisfied and copes with his life and the therapy which is being administered. QL is influenced by physical, psychological and social factors as well as by time. During the individual course of a malignant disease and its therapy, changes in QL need to be assessed with the appropriate "measuring instruments". Once the quality of the data has been assured, comparisons can be made with respect to the influence of alternative therapies on QL. This is where "promotional" information from the pharmaceutical industry can be emphasized; QL degenerates to a useless term, however, if it is used indiscriminately and without proper validation of the data. A new concept, i.e. QUALYs (= Quality Adjusted Life Years), has originated in the USA, which attempts to add a dimension of currency value to the term LQ.


[The effect of starvation on mast cell functional activity and on the content of microsomal cytochromes in the liver].

Histamine secretion and ATP levels were measured in mast cells from the abdominal and pleural cavities of Wistar rat males (250-300 g) which had been starving for 2-4 days. Food deprivation had no effect on histamine levels and secretion in exposure to substance 48/80. ATP concentration got reduced from starvation day 2. Food deprivation for 2 days led to decline in the content of cytochromes P-450, P-450B, in the ratio P-450B/P-450L, but had no effect on rat liver cytochrome B5. 4-day starvation did not entail further changes.


Evaluation of expanded polytetrafluoroethylene (e-PTFE) and autogenous fascia lata in frontalis suspension. A comparative clinical study.

Ptosis palpebrae superioris is mainly treated by frontalis suspension with autogenous fascia lata. In this study we evaluated the potential of an expanded polytetrafluoroethylene (ePTFE) soft tissue patch to function as a frontalis sling in 13 patients with ptosis palpebrae superioris. Functional and cosmetic results and the satisfaction of the patients were scored positive or negative and statistically compared with 13 patients in whom fascia lata was used. The mean follow-up was ten months. Although no complications occurred in either group the patients in the ePTFE group were less satisfied (p < 0.01) and showed inferior functional (p < 0.05) and cosmetic results (p < 0.01) as compared to the fascia lata group. Histological analysis of an explanted piece of ePTFE revealed that tissue ingrowth into the patch was lacking, which might have led to poor anchorage and subsequently a recurrence of the ptosis. We believe that ePTFE is not the material of first choice in frontalis suspension.


Long-term experiences with the two-stage palatoplasty with regard to the development of maxillary arch.

From our data on 46 patients operated on by one surgeon in 1961 and 1962 who were regularly examined up to adulthood, it may be concluded that the growth results of the two-stage palatoplasty, when the second stage is done before the patient is 7 years of age, are almost identical to the results of one-stage palate repair performed at the age of 3 to 4 years. However, a greater orthodontic effort is needed to achieve a properly aligned dentoalveolar arch with the two-stage palatoplasty.


An extensive defect on the tibia covered by a free cross flap using m. latissimus dorsi.

Using a clinical case, the authors demonstrate the technique of covering an extensive tibial defect unmanageable, due to blood supply to the limb and extent of the defect, by conventional methods. The required amount of tissue was provided by a flap obtained from m. latissimus dorst transferred to the site of defect as a free cross flap.


Extracorporeal tissue transfer for salvage replantation.

The author reports the use of extracorporeal tissue transfer technique for salvaging severed digits of the hand which are unfit for replantation. The technique of extracorporeal tissue transfer has been reported by us earlier. In this paper its use has been extended for salvaging five non replantable digits in two cases.


The neck collar--a treatment of hypertrophic burn scars of the neck region. A clinical study.

Hypertrophic burn scars of the neck region are disfiguring and cause functional disturbances such as itchiness, tenderness, blistering or even contractures. Since 1970 the Burns Unit in Copenhagen has treated hypertrophic scars of the neck with a special soft neck collar as a routine. This pressure treatment is controlled in the Jobst day-clinic by a team consisting of a surgeon, physiotherapist, occupational therapist and a nurse. The clinical results of 17 cases are reported and commented on.


Growth and development of the face in complete unilateral cleft lip and palate during prepubertal and pubertal periods.

Roentgencephalometry was used for the assessment of the growth and development of the face during prepubertal and pubertal periods in children with complete unilateral cleft lip and palate treated with the same methods. The first series included 16 boys and 15 girls, the second series consisted of 15 boys and 15 girls examined repeatedly at the beginning and at the end of the follow-up period. There were no definite differences in the growth rate of the face between the two periods of age. Therefore, the deterioration of overjet during puberty could be due to the depletion of the compensation and adaptation mechanisms after the previous orthodontic treatment rather than to the enhanced growth rate. As compared to other facial dimensions a lower growth rate showed during both periods the length of the anterior base, the depth of the maxilla and in particular the height of the upper lip. Developmental changes of variables of shape and position proceeded similarly during both periods of age. An exception represented the rotation of the mandible and the inclination of upper incisors. During the prepubertal period the lower jaw showed a very slight posterior rotation, while during puberty an anterior growth rotation was present. A rapid improvement of the proclination of upper incisors was attained only during the prepubertal period. It was accompanied by an improvement of overjet. However, during the period of puberty there was a renewed deterioration of overjet. A marked retrusion of the maxilla developed already in the prepubertal period. During both periods occurred an identical impairment of sagittal jaw relations and of the upper lip prominence, accompanied by a flattening of the facial profile and reduction of the nasolabial angle. The prominence of the nose increased, the angle of the cranial base remained unchanged. Intersexual differences were not demonstrated in any studied characteristics.


Ethical problems in burn injury in the past decade.

As a result of the increasing numbers of disasters and constant advances in medical care technology allowing survival of critically ill patients by using an interdisciplinary approach to comprehensive therapy, the past decade has seen the emergence of ethical problems related to burn injury. The study discusses the issue of sorting out patients from the point of view of providing emergency care and transport as well as from the point of view of psychologic care and palliative therapy. The quality of life and prolonged suffering of elderly individuals is another major issue as well as, last but not least, ethical and moral obstacles encountered in clinical research carried out in the critically ill where the potential of misuse is greater than in other groups of patients. The role played by ethical committees and by integrity of the research project coordinator are highlighted.


The use of ozonotherapy in the nose correction operations.

Positive results of application of ozonotherapy in rhinoplasty are described. 25 patients were administered ozone intravenously for 3 days prior to operation and days post-operatively, resulting in a significant reduction in post-operative complications duration as compared with the control group consisting of 20 patients who were not administered ozonotherapy.


[Chromosomal determinant of Streptomyces hygroscopicus with positive influence on resistance to bialaphos controlled by the bar gene].

Streptomyces hygroscopicus ATCC21705 is an organism producing bialaphos, a promising ecologically safe antibiotic of peptide nature. An unknown determinant was cloned within the 1.4-kb SsH fragment of chromosomal DNA of S. hygroscopicus. Its presence along with the bar gene in the hybrid plasmid PVGB21 increased expression of the phenotype resistance to bialaphos in the recipient strain of S. lividans TK 64. Plasmid PVG B22 carrying the isolated SsH fragment did not provide resistance to the antibiotic. Transcription from the cloned fragment was revealed by RNA-DNA hybridization. It was shown that the cloned determinant had a positive action on bar gene transcription correlating with a significant increase in the activity of N-acetyltransferase in the mycelium of the transformant pVG B21.


RNase protection assays and RNA gel blots: a direct comparison of sensitivity.

RNase protection assays are commonly thought to be a more sensitive means of detecting and quantitating specific mRNAs than are RNA gel blots (Northern blots). We have directly compared the sensitivity of these two approaches by assaying for known amounts of in vitro synthesized beta-glucuronidase mRNA. With the probes and protocols employed here, the ability to detect a specific mRNA was similar whether RNase protection or RNA gel blot analyses were performed.


Direct reverse transcription--polymerase chain reaction from whole blood without RNA extraction.

A simple, fast, efficient method of direct reverse transcription-polymerase chain reaction (RT-PCR) from whole blood without RNA extraction is described. After the erythrocytes in a few microliters of fresh blood were destroyed and removed, the total RNA was released from the leukocytes by means of water treated with diethyl pyrocarbonate. The solution was used as RNA template and directly introduced in the RT-PCR. Following this method, which is called direct RT-PCR, three different cDNA fragments were successfully amplified with three different pairs of primers, respectively. Meanwhile, the direct RT-PCR was compared with the RT-PCR from the extracted total RNA template. The results showed no obvious difference.


Three-dimensional structures of the human alpha 2-macroglobulin-methylamine and chymotrypsin complexes.

The three-dimensional structures of chymotrypsin- and methylamine-treated negatively stained human alpha 2-macroglobulin have been determined by weighted back projection from electron microscope data. Projections of the reconstructions show good concordance with two-dimensional averages of both stained and frozen-hydrated molecules. The reconstructions reveal that the H-shaped front projection of the molecule is related to the smaller ellipsoidal end view by a rotation of 90 degrees about the crossbar (minor axis) of the H. This finding is in agreement with tilt studies. The reconstruction of the alpha 2-macroglobulin-methylamine reveals an hour-glass shaped void which is filled by the two proteinase molecules in the reconstruction of alpha 2-macroglobulin-chymotrypsin. Protein plugs which appear to block the exterior entrances to the cavity may function to prevent access of proteins to the encapsulated proteinase and serve to block its escape. Extensive thresholding of each reconstruction leaves a "backbone" consisting of two side-by-side rod-like structures, suggesting that this is the arrangement of the two protomeric units which form the molecule. Both structures show some departure from the expected symmetry. The asymmetries are robust features of the reconstructions and may reflect structurally asymmetric features of the transformation from the native to the chymotrypsin-treated form of the molecule.


[A method for isolating a complex epoxide-containing antigen and its use for the diagnosis of allergic diseases].

Specific complex antigen, based on human serum albumin and epichlorhydrine was proved to be useful in the diagnosis of individual hypersensitivity to the epoxide pitches and epichlorhydrine. Lymphocytes' blast transformation with 3H-timidine was proved to be the most sensitive reaction in the use of specific complex antigen during the diagnosis. The stated immune reaction can be used for the diagnosis of allergy before the clinical manifestations.


Frequency of common colds and serum levels of sICAM-1 (CD54), sLFA-3 (CD58) and sIL-2R (CD25).

Epidemiologic case-control studies have suggested an inverse relationship between past medical history of frequency of common colds and subsequent cancer risk for various sites. One hypothetical explanation for this finding may be that there are unknown differences in immune responsiveness between patients developing cancer and healthy individuals. The present study examines the relationship between the frequency of common colds and a) plasma levels of sICAM-1/CD54, sLFA-3/CD58 and sIL-2R/CD25 which are believed to modulate activation of immune responses, and b) cell-mediated immunity in vivo as determined by Multitest Mérieux. The investigation was conducted as a correlation study amongst a healthy group of individuals from the general population in Germany. We found a statistically significant inverse relationship between frequency of common colds and levels of sCD58 and, partially, sCD54. No association was found between levels of sCD25 and results of Multitest Mérieux.


[The comparative characteristics of the cytochemical indices of the peripheral blood neutrophils in the separate administration of organochlorine and organophosphate pesticides against a background of dynamic loading with different intensities].

Cytochemical parameters of peripheral neutrophils were studied in the experimental pattern of combined action of chlorine and phosphorus containing organic pesticides and different levels of dynamic physical strain. Studies revealed the protective effect of the minimal physical strain and aggravating effect of the moderate and maximal strain leading to the depressed aerobic oxidation and increased anaerobic glycolysis.


Endotoxin tolerance: regulation of cytokine production and cellular changes in response to endotoxin application in cancer patients.

Endotoxin (lipopolysaccharide, LPS) has the property of inducing tolerance to its own biological effects. This phenomenon has been extensively studied in animal models but only few studies exist on the regulation in humans. Here we describe experiments designed to determine the cytokine regulation and cellular changes in humans during induction of LPS tolerance after repeated LPS injections. Intravenous administration of purified LPS Salmonella abortus equi to cancer patients induces high amounts of circulating tumor necrosis factor-alpha (TNF-alpha), interleukin-6 (IL-6), interleukin-8 (IL-8), granulocyte colony-stimulating factor (G-CSF), and macrophage colony-stimulating factor (M-CSF). Repeated injections of LPS at daily intervals resulted in a marked downregulation of the cytokine response and in the case of TNF-alpha, IL-8, G-CSF, and M-CSF the cytokine response was reduced to baseline levels. In contrast, significant increases in serum IL-6 were detected up to day 5 of repeated LPS injections. Hematological changes included transient decreases in WBCs affecting granulocytes, monocytes, and lymphocytes, followed by a marked granulocytosis. The drop in WBCs remained unaltered throughout the 5 day course of repeated LPS injections whereas the granulocyte overshoot recovery diminished gradually. When PBMCs of the cancer patients were restimulated ex vivo a marked enhancement of the capacity to produce TNF-alpha, IL-113, and IL-6 occurred, which is in contrast to the decreasing TNF-alpha serum levels obtained in vivo. In parallel, a shift in monocyte subpopulations from CD14+/CD16- to CD14+/CD16+ cells was observed. The data provide evidence that different mechanisms are implicated in the cytokine downregulation following repeated LPS injections to cancer patients. Furthermore, PBMCs from LPS tolerant patients do not demonstrate a reduction in their capacity to produce cytokines.


A tiaramide derivative, 5-chloro-3-(4-hydroxypiperadinocarbonylmethyl)benzothiazoline-2-one (HPR-611), a potent inhibitor of anaphylactic chemical mediator release--a distinctive feature from disodium cromoglycate.

Effects of a tiaramide derivative, 5-chloro-3-(4-hydroxypiperadinocarbonylmethyl)benzothiazoline++ +-2-one (HPR-611), on anaphylactic chemical mediator release from rat peritoneal exudate cells (RPEC), guinea pig lung fragments (GPLF) and human lung fragments (HLF) were investigated in comparison with those of tiaramide and disodium cromoglycate (DSCG). HPR-611 at 10(-6) - 10(-4) g/ml showed a concentration-dependent inhibition of the histamine release from RPEC regardless of its pretreatment time. Tiaramide also inhibited the release with slightly less potency than HPR-611. The treatment of DSCG 1 min before antigen challenge markedly prevented the release but the inhibitory potency was clearly deteriorated by prolongation of the pretreatment time. Tiaramide tended to influence the anaphylactic release of histamine from GPLF with only 20% inhibition of the release at either 10(-5) or 10(-4) g/ml, whereas HPR-611 at 10(-5) and 10(-4) g/ml significantly suppressed the release in a concentration-dependent fashion. DSCG was not effective on that even at higher concentrations. Anaphylactic release of not only histamine but also immunoreactive leukotriene B4 (i-LTB4) and i-LTC4 from HLF was markedly inhibited by 10(-8) - 10(-4) g/ml of HPR-611. Tiaramide inhibited the release to a somewhat less extent than HPR-611, while nominal or no inhibitions by DSCG were found. From these results, it is clearly apparent that anti-allergic actions of HPR-611 are quite different from those of DSCG.


Prostate adenocarcinoma using Gleason scores correlates with prostate-specific antigen and prostate acid phosphatase measurements.

To evaluate a relationship between Gleason scores of histopathology of prostate carcinoma and concurrent serum prostate-specific antigen (PSA) and prostate acid phosphatase (PAP) values, 65 men with prostate carcinoma were studied. These patients' cumulative Gleason scores were obtained by totaling the primary and secondary patterns, resulting in two groups: 42 patients received high (6-10) and 23 received low (2-5) Gleason scores. Serum PSA and PAP values were measured by radioimmunometric assay 1 to 7 days before surgical procedures or biopsy for prostate carcinoma. Mean serum PSA for patients in the high Gleason score group was 134.39 ng/mL (normal range: 0 to 4), and the mean serum PSA for patients in the low Gleason score group was 23.62 ng/mL. Mean serum PAP for patients with high scores was 28.08 ng/mL (normal range: 0 to 5), and the mean serum PAP for patients with low scores was 18.19 ng/mL. Patients with high Gleason scores showed significantly greater elevation of serum PSA than those with low Gleason scores (P = .047), using two samples to test for groups having unequal variants. Prostate acid phosphatase levels of patients with high scores were not significantly higher than the levels in patients with low scores (P = .60). These results indicate that PSA levels but not PAP levels correlate with Gleason scores.


Effect of neutralizing monoclonal antibodies on Hantaan virus infection of the macrophage P388D1 cell line.

The effect of neutralizing monoclonal antibodies (N MAbs) on Hantaan virus infection of macrophages was investigated using P388D1 cells, a murine macrophage cell line. MAbs to the G1 protein (G1b) and the G2 protein (G2a and G2c) neutralized viral infectivity in P388D1 cells. N MAbs to G1b showed much higher neutralizing potency than those to G2a and G2c. With each N MAbs, two distinct effects were observed: neutralization of viral infectivity occurring at high concentrations and enhancement of that at low concentrations. Non-neutralizing MAbs, on the other hand, showed only enhancement of viral infectivity even at high concentrations without any inhibitory effects. The Fab fragments of N MAbs showed neither neutralizing nor enhancing activities. However, when the virus-Fab complexes were reacted with the anti-Fab antibodies, both neutralization and enhancement of viral infectivity were restored depending on the dose of Fab fragments. These results indicate that Hantaan virus infection of P388D1 cells is mediated by the Fc portion of the antibodies and neutralization is dependent on the concentration of N antibodies bound bivalently to the neutralization site on the virion.


A pilot study of ChlOPhEPP-B--a weekly regimen for patients with untreated advanced or recurrent Hodgkin's disease.

Twenty-five patients with either previously untreated advanced Hodgkin's disease (HD) or with relapsed disease were treated with an intensive weekly chemotherapy regimen (ChlOPhEPP-B) in which the myelosuppressive combinations of epirubicin and chlorambucil, and procarbazine and etoposide alternated at two-week intervals, interspersed with vincristine and prednisolone and then vincristine, prednisolone and bleomycin in a four-week cycle. Of the previously untreated patients (n = 14), 12 (86 per cent) showed a complete response (CR); in the previously treated patients (n = 11), CR was achieved in five (46 per cent). Toxicity was predictable and not evidently more severe than with more standard regimens. Neutropenia leading to treatment delay and/or dose reduction was seen in 11 patients (44 per cent) but severe neutropenia (WHO grade 4) was documented in only six patients (24 per cent) for a cumulative total of nine treatment weeks (1.8 per cent of the total treatment time). Dose reductions for neutropenia were applied in relation to 11 of the 379 weekly treatment points (2.9 per cent). Delays in treatment, for any reason, averaged 3.65 weeks per patient. The study demonstrated the feasibility of a more intensive weekly multidrug schedule. This approach may be applicable in the treatment of advanced HD and represents a further option in the strategy of treating relapsed disease.


Differential distribution of a carbohydrate epitope (Y) on human salivary gland cell membranes.

Monoclonal antibody 303 (mAb 303) reacts with the high molecular weight agglutinin present in human saliva. Its reactivity is periodate sensitive, and it has been shown to recognize the Y epitope. Immunogold labeling of thin sections of human parotid and submandibular glands with mAb 303 showed reactivity in secretory granules of serous acinar, intercalated and striated duct cells (Takano et al., 1988). We now report that the apical and basolateral membranes of salivary acinar and duct cells are labeled by mAb 303, but not myoepithelial cells, endothelial cells and other mesenchymal cells. Gold particles were confined to acinar and duct cell membranes even when myoepithelial cells were directly adjacent, suggesting that the epitope resides on a membrane glycoprotein and that the label does not represent secreted agglutinin bound to the cell surface. Although myoepithelial cells are thought to differentiate from epithelial stem cells, the present results indicate that substantial compositional differences exist between the membranes of myoepithelial cells and other salivary parenchymal cells. Earlier studies also showed that mAb 303 labels normal pancreatic acinar cells and certain salivary (pleomorphic adenoma) and mammary (lactating adenoma) tumors (Bogert et al., 1988). This antibody thus may be a useful reagent for characterizing the origin of exocrine gland-derived cell cultures and neoplastic cells. Further, localization studies may provide insight into the role of the Lewis blood group-related epitope in secretory cells.


Viral inhibition of the interferon system.

In response to interferon (IFN), cells develop an antiviral state in which the replication of a wide spectrum of RNA and DNA viruses is inhibited. Viruses have evolved a variety of mechanisms to inhibit the production and action of the interferons. Interferon action may be blocked by inhibition of the post-receptor signalling pathway, which prevents the expression of a number of proteins with antiviral properties. Other viruses prevent the action of specific, interferon-induced antiviral systems. In particular, the action of the dsRNA-dependent protein kinase (DAI) is inhibited by a variety of different viruses, indicating the fundamental importance of this enzyme to the antiviral response.


Specific interactions of pancreatic amylase at acidic pH. Amylase and the major protein of the zymogen granule membrane (GP-2) bind to immobilized or polymerized amylase.

Regulated secretory proteins are thought to be sorted in the trans-Golgi network towards the secretory granule via acidic aggregation. In the exocrine pancreas, amylase is one of the major zymogens. It is a basic protein of pI 8.6 and does not precipitate in acidic conditions. To identify the mechanism by which amylase aggregates in the acidic cisternae of the pancreatic trans-Golgi network, we have developed an in vitro model in which amylase was fixed to plastic microtiter plates. The fixed amylase was probed with two ligands: amylase itself and GP-2, the major protein of the zymogen granule membrane. Biotinylated amylase bound to fixed amylase in a strict pH-dependent manner with optimal binding between pH 5.0 and 5.7. The affinity of binding was in the nanogram range (Kd approximately 20.0 ng/mL) at pH 5.5. Acid binding of amylase was not reversible by incubation at neutral pH, nor could it be displaced by native amylase. GP-2 binding to fixed amylase was also pH dependent with optimal binding between pH 5.0 and 5.7. As for amylase, it was not reversible by incubation at neutral pH. GP-2 binding sites on fixed amylase appeared to be different from those of biotinylated amylase. While native and biotinylated amylase did not bind to GP-2, polymerized amylase precipitated GP-2 at acidic pH. Taken together these data suggest that slight modifications are sufficient to reveal on the amylase molecule binding sites for GP-2 and for amylase itself. These new binding capacities acquired at acidic pH could be involved in the cascade of reactions that lead to the in vivo formation of the immature secretory granule.


The production of hybridoma cell line secreting monoclonal antibodies against Xanthomonas campestris pv. oryzae and its application in the classification of strains.

By the fusion of mouse myeloma cells (SP2/0-Ag14) and spleen cells derived from BALB/c mice immunized with the preparation of Xanthomonas campestris pv. oryzae Ks-6-6, Os-213, Yz-32 and Yz-24, we obtained 12 hybridoma cell lines secreting monoclonal antibodies. None of the McAbs cross-reacted with the other varieties of plant pathogenetic and non-pathogenetic bacteria. The McAbs could distinguish three variant serotypes of strains. Antibody titers of ascites were about 1:10(3)-1:10(6) when measured by ELISA method. The McAbs could differentiate 6 epitopes. Based on the epitopes, the 63 strains of X. campestris pv. oryzae we collected were grouped into nine groups.


Overexpression of rnc gene and purification of RNaseIII.

The reason for low content of RNaseIII in E. coli is that RNaseIII has a negative feedback action on its own synthesis by cutting off the transcripts of its own gene, rnc, at 5'-terminal. On this basis, the scheme for overproduction of RNaseIII was designed. The 5'-flanking sequence of rnc gene transcribing to form a secondary structure which could be degraded by RNaseIII was removed, and the whole coding sequence, including the translational initiation signal, was reserved and put under the control of lambda PL promoter. The constructed plasmid pCR21, which contain the recombined rnc gene, overproduced RNaseIII, which covered over 65% of the total cell protein and formed inclusive bodies in E. coli after induction at 42 degrees C. By using the characteristics of solubility of the protein, electrophoretic pure RNaseIII was obtained with a simple procedure, including lysis of the bacterial cells, washing precipitates of the lysate repeatedly at low temperature and low salt concentration, and dissolving and passing through Q-Sepharose FF column in high salt concentration at room temperature. The yield of purified RNaseIII was 10-12 mg per 100 ml culture, and lambda sib transcripts were cut at special sites. RNaseIII possessing the activity of binding ATP is reported.


Pharmacological doses of insulin equalize insulin receptor phosphotyrosine content but not tyrosine kinase activity in plasmalemmal and endosomal membranes.

Following insulin administration to intact rats, the insulin receptor kinase activity of subsequently isolated cell fractions was significantly augmented. Of interest was the observation that the endosomal insulin receptor tyrosine kinase displayed four- to six-fold greater autophosphorylation activity than that of plasma membrane. Surprisingly, the endosomal insulin receptor tyrosine kinase displayed a decrease in beta-subunit phosphotyrosine content compared with that seen in the plasma membrane. These observations prompted the suggestion that insulin receptor tyrosine kinase phosphotyrosine dephosphorylation mediated by an endosome-specific phosphotyrosine phosphatase(s) yields activation of the endosomal insulin receptor tyrosine kinase. In a previous study we examined the effect of subsaturating doses of injected insulin. In this work we evaluated insulin receptor tyrosine kinase activity and phosphotyrosine content in plasma membrane and endosomes after a receptor-saturating pharmacological dose of insulin (150 micrograms/100 g body weight). At this dose the phosphotyrosine content per receptor was reduced compared with that seen earlier at insulin doses of 1.5 and 15 micrograms/100 g body weight. Endosomal insulin receptor tyrosine kinase was greater than that seen at the lower nonsaturating insulin doses. Furthermore, endosomal insulin receptor tyrosine kinase activity exceeded that of the plasma membrane, despite retaining about the same phosphotyrosine content per receptor. These data are consistent with the view that insulin receptor tyrosine kinase activity may be regulated by a particular pattern of phosphotyrosine content on the beta-subunit wherein both activating and inhibitory phosphotyrosine residues play a role.


Modulation of cytokeratin and actin gene expression and fibrillar organization in cultured rat hepatocytes.

Intermediate filaments of rat hepatocytes are composed of cytokeratins 8 and 18 (CK8 and CK18, respectively). Recent work from our laboratory has indicated a close relationship between the synthesis of these cytokeratins, their organization into intermediate filaments, and the promotion of growth and differentiation of cultured rat hepatocytes by insulin, epidermal growth factor, and dexamethasone. In the present study, we examined the mRNA expression, level of protein synthesis, and fibrillar distribution of cytokeratins 8 and 18 and actin in hepatocytes, isolated from normal and dexamethasone-injected rats and cultured as monolayers or spheroids in the presence of insulin, or from normal rat hepatocytes, cultured as monolayers in the presence of dexamethasone, insulin, and dimethyl sulfoxide. The CK8 mRNA level was lower in hepatocytes isolated from noninjected rats and cultured as either monolayers or spheroids, than in those from dexamethasone-injected rats. However, the CK18 mRNA level varied in a manner that was different from that of CK8 mRNA, showing that the modes of expression of the two genes were independent. The various changes in hepatocyte culture conditions led to variations in albumin mRNA levels that largely followed those observed in CK8 mRNA levels. In the case of actin, the amount of mRNAs varied from relatively high levels in hepatocyte monolayers to extremely low levels in hepatocyte spheroids, even though in both cases the cells were isolated from dexamethasone-injected rats. These changes in mRNA levels did not necessarily correlate with changes in the synthesis of cytokeratins 8 and 18, and actin. Changes in culture conditions induced a major reorganization in the distribution of cytokeratin intermediate filaments and actin filament between the region near the surface membrane and the cytoplasm. The most divergent patterns in cytokeratin intermediate filaments and actin filament distributions were observed between hepatocytes cultured as spheroidal aggregates and as monolayers in the presence of dimethyl sulfoxide. The former condition resulted in patterns of cytokeratin and actin gene expression and fibrillar organization that best matched those in situ. In the latter condition, inappropriate patterns were obtained, in spite of the fact that dimethyl sulfoxide treated hepatocytes are known to exhibit survival and functional activities equivalent to that of hepatocyte spheroids. These results demonstrate for the first time that the survival and functional activity (i.e., albumin production) of rat hepatocytes in vitro is not necessarily correlated with a particular pattern of cytokeratin and actin gene expression and fibrillar arrangement.


Isolation of 2 novel RFLP markers and their localization at 2q35 by microdissection and subsequent enzymatic amplification.

We previously constructed a chromosome 2q-specific genomic library and isolated a number of microclones. In the present study, we first analyzed with Southern hybridization whether any of the microclones represent restriction fragment length polymorphisms (RFLPs), and then tried to map RFLP markers physically, using the recently developed chromosome microdissection/enzymatic amplification method. Of 13 clones analyzed, two were RFLP markers; a clone, pM2C83, showed a four-allele MspI RFLP, and the other, pM2C8, a two-allele RsaI RFLP. In order to assign the two polymorphic markers, two chromosomal segments, 2q32-q35 and 2q35-qter, on the chromosome 2 from a karyotypically normal person were microdissected, and the DNA from each segment was amplified with the polymerase chain reaction (PCR) using marker sequence-specific primers. With this method, both of the clones were assigned to 2q35. These two RFLP markers must be useful for linkage analysis of genetic diseases whose loci are at around 2q35.


Parent-specific expression of a human keratin 18/beta-galactosidase fusion gene in transgenic mice.

Insertion of a human keratin 18 (K18)-bacterial beta-galactosidase (LacZ) fusion gene into mice has led to a unique transgenic line in which expression of the transgene is subject to unusual germ line-specific, genomic imprinting effects. Fetal expression of the LacZ reporter gene depends on the gender of the transmitting parent, with appropriate expression in liver after maternal inheritance, and ectopic expression in retina and mesodermal tissues after paternal inheritance. This tissue-specific imprinting pattern is superimposed upon a basic expression pattern which is unaffected by parental inheritance. Insertion of the transgene has led to a recessive-lethal phenotype, with no parent-of-origin effects on viability, suggesting that the transgene has not inserted into an imprinted region of the genome. HpaII and HhaI methylation sensitive restriction sites within the bacterial LacZ reporter gene are completely methylated when activity of the maternally inherited transgene is detected in the fetal liver, and not methylated when the paternally inherited transgene is silent. Thus DNA methylation of LacZ is correlated with maternal inheritance and may be implicated in the genomic imprinting mechanism as others have suggested. However, in contrast to the commonly found correlation of expression and low DNA methylation, the LacZ gene was expressed in fetal liver when fully methylated. This result may imply the existence of negative regulatory activities that recognize the unmethylated LacZ gene.


Trying to overcome residual disease after bone marrow transplantation for hematologic malignancies.

Relapse after marrow transplantation occurs despite total body irradiation and high doses of chemotherapy used to prepare the recipient. Recurrence rate is particularly high after autologous transplantation. In order to eliminate minimal residual disease persistent after marrow grafting, immunotherapy is currently being explored as a treatment modality that is non-cross-reactive with radiation and chemotherapy. This review describes first clinical results on the use of interferons and interleukin-2 in marrow transplant recipients as well as attempts to induce graft-versus-leukemia effects.


[Slowly progressive dysarthria and impaired language function--a case report].

A 68-year-old right-handed woman was admitted to Tokyo Metropolitan Geriatric Hospital because of slowly progressive dysarthria and writing disability over 2-year period. On admission, severe dysarthria was observed, but no dysphagia. The dysarthria mostly resembled a type of pseudobulbar palsy, although it was associated with effortful speech production. An oro-facial apraxia was also found. She could name objects, and could understand spoken words correctly. Examination using the Western Aphasia Battery showed diminution of word fluency, impaired repetition and perseveration and writing errors. On the Wechsler Adult Intelligence Scale-R verbal IQ was 100 and performance IQ was 87. These scores did not suggest any significant degree of general intellectual deterioration. Wisconsin card sorting test disclosed mild frontal dysfunction. Magnetic resonance imaging showed cortical atrophy in the bilateral frontal and temporal lobes. Measurements of regional cerebral metabolic rate by 18F-FDG-PET demonstrated decreased uptake in the latero-dorso-inferior area of the bilateral frontal lobes, especially on the left side. The present case showed slowly progressive dysarthria and progressive aphasia without generalized dementia, and without typical aphasia. These symptoms are speculated to be related to the atrophy in the bilateral frontal and temporal lobes shown by MRI and the decreased metabolic rate in the left dominant bilateral frontal lobes on PET study. The pathologic process responsible for these lesions remains obscure.


Immunoperoxidase labelling of previously stained tissue sections: application in oral histopathology.

We have investigated by the avidin-biotin peroxidase technique a wide variety of recognized cellular antigens in paraffin sections of both normal and pathological material which have been previously stained with hematoxylin and eosin. The intensity of peroxidase reaction was adequate to permit the diagnosis and the background of the immunoreaction minimal. This technique therefore, appears useful either for retrospective analysis of cellular antigens or when limited unstained material is available.


[Epidermoid carcinoma of the oral cavity: a histopathological study of 85 cases].

In 85 cases of intraoral squamous cell carcinoma we have carried out a descriptive and statistical analysis of 8 parameters or morphological aspects: structural growth pattern, degree of keratinization, differentiation or similarity, nuclear pleomorphism, frequency of mitoses, inflammatory response, number of eosinophils and vascular invasion. We have found that in 65% of the cases there was a cord type growth pattern, and 51% evidenced a maximum degree of keratinization. The neoplastic cells were characterized by a moderate differentiation and pleomorphism, 69% and 73% respectively, there being 1-3 mitoses/8 fields in 54% of the cases. We did not find very marked differences between degrees 1, 2, and 3 of inflammatory response (39%, 25%, 33% respectively). We observed vascular invasion in only 8.5% of the cases. The average value of intratumoral and peritumoral eosinophils was 18 and 37.


[Comparison between nuclear DNA and histologic grade of prostatic carcinoma].

DNA nuclear ploidy determined by flow cytometry was evaluated from prostate tissue in 51 patients with prostatic cancer who had undergone radical prostatectomy. DNA ploidy pattern was diploid in 46% and aneuploid in 54% of tumors. DNA ploidy was compared to histological tumor grading. 92 Aneuploidy was found in 0% of the tumors with Gleason score between 2 and 4 in 62% between 5 and 7 and in 50% between 8 and 10. Our results suggest there is no relationship between the two parameters.


Conversion of Mobitz type II AV block to 1:1 AV conduction by premature ventricular beats.

Preliminary experiments in a canine model of Mobitz type II atrioventricular (AV) block showed improvement of conduction after premature ventricular beats. In this investigation, the authors studied the mechanism(s) responsible for this response. In vivo studies were performed in 16 anesthetized dogs. Block was induced by ischemia after septal artery occlusion or by mechanical trauma. Two pairs of plunge electrodes were inserted in the proximal and distal His bundle. An electrode catheter was positioned at the level of the aortic root to provide an overall view of His bundle activation. Bipolar pacing was performed from the high right atrium, right ventricular outflow tract, and proximal and distal His bundle. Infra-nodal 2:1 AV block was consistently induced at an atrial rate of 238 +/- 21 beats/min. In 15 dogs a narrow time window (10-60 ms; mean, 32 +/- 6 ms) was found during which premature beats resulted in transient (2-11 beats; n = 9) or persistent (n = 8) restoration of 1:1 AV conduction. Retrograde penetration of the site of block, that is, Hb, was found even when the anterograde impulse was blocked, demonstrating the asymmetric nature of anterograde versus retrograde conduction. In vitro studies were performed in the same hearts. Intracellular recordings were obtained in the damaged His bundle and proximal right bundle. The site of block showed frequent displacements along the bundle. The introduction of a retrograde stimulus during 2:1 block restored 1:1 anterograde conduction.(ABSTRACT TRUNCATED AT 250 WORDS)


Endothelin-1 inhibits the vasodilation induced by substance P in isolated porcine ophthalmic artery.

The vasoconstrictor effect of endothelin-1 is long-lasting and slow in onset, suggesting that this peptide may be involved with long-term vascular responses. Substance P, a quick-acting, vasodilatory tachykinin present in sensory nerves, may be released by trigeminal fibres and thereby participate in the regulation of vascular tone in the head. Given the potential role of vasoregulatory peptides in the ocular-forehead circulation in the pathophysiology of vascular headaches, the interactions between substance P and endothelin-1 over time were studied in isolated porcine ophthalmic artery. Substance P-induced relaxation of precontracted artery segments was significantly diminished by endothelin-1, despite the fact that endothelin-1 itself had no further significant contractile effect on these arteries. It is also noteworthy that the observed reduction of substance P-induced relaxation occurred much more rapidly than the onset of endothelin-1-induced contraction in relaxed porcine ophthalmic artery. The results suggest an inhibitory synergism between endothelin-1 and substance P in this vascular bed.


Structure of G-CSF: significance of the sugar chain.

1. The carbohydrate chain protects rhG-CSF from polymerization and/or conformational alterations associated with physicochemical changes, elevation of pH or temperature fluctuations. 2. The carbohydrate chain of rhG-CSF prevents loss of its biological activity in normal human serum by inhibiting proteinase activity. 3. These facts indicate that the carbohydrate chain of rhG-CSF has a markedly important role in maintaining the stability of the protein itself as well as in effecting the exertion of its biological activity.


G-CSF mobilised progenitor cells in autologous transplantation: in vitro and in vivo aspects.

G-CSF administration leads to significant elevation in the levels of circulating progenitor cells. Infusion of these cells after high-dose chemotherapy is associated with accelerated platelet engraftment that has a considerable impact in shortening thrombocytopenia and reducing need for platelet transfusions.


Inward rectification of acetylcholine-elicited currents in rat phaeochromocytoma cells.

1. Currents elicited by acetylcholine (ACh) were studied in the rat phaeochromocytoma cell line, PC12, using patch-clamp techniques. 2. Whole-cell ACh-elicited currents are inwardly rectifying and intracellular Mg2+ can play a role in determining the extent of whole-cell current rectification. Increasing the intracellular Mg2+ concentration, [Mg2+]i, augmented the rectification. The effects of increased [Mg2+]i on the whole-cell current can be explained by the block of receptor channels by Mg2+. 3. In the nominal absence of internal divalent cations, however, a substantial degree of rectification remains. This rectification is probably not due to divalent cations, as buffering the external Mg2+ concentration to 50 microM and the internal concentration to nominally 0 Mg2+ did not reduce the rectification. The remanent rectification was not due to block by the main permeant cation, Na+. Using K+ or Cs+ as the main monovalent cation inside the cell did not diminish the rectification. Neither replacing the pH buffer, HEPES, with phosphate buffer nor increasing the intracellular pH removed the rectification. 4. For ACh receptor channels in excised patches, the voltage dependence of the probability of being open (Popen) stemmed mainly from the voltage dependence of the channel burst duration. The channel opening rate was relatively voltage independent. The weak voltage dependence displayed by the channel burst duration was insufficient to account for the reduced whole-cell outward current at positive potentials. The mean burst duration of the channel did not have a simple logarithmic relationship with voltage. 5. In the absence of intracellular Mg2+, the instantaneous current-voltage relationship for whole-cell currents was linear suggesting that the I-V relationship of single channels in perfused cells is linear and does not contribute to the rectification of the whole-cell current. 6. In perfused cells, receptor channels had a low steady-state probability of being open at positive potentials compared to channels in excised patches. Voltage jumps to positive potentials revealed a process in perfused cells which could account for the low Popen. Relaxations of agonist-induced current at +40 mV had a large, exponentially decaying component that quickly closed channels (rate constant, tau, approximately 400 microseconds). The mechanism responsible for this decay could explain the rectification that remains in the absence of intracellular divalent cations.(ABSTRACT TRUNCATED AT 400 WORDS)


Effect of nitro-L-arginine on endothelium-dependent hyperpolarizations and relaxations of pig coronary arteries.

1. Endothelium-dependent relaxation is caused by an endothelium-derived relaxing factor (EDRF) identified as nitric oxide (NO). Our objective was to test whether one or several distinct endothelium-dependent relaxing factors exist. 2. In pig coronary arteries, a hyperpolarization accompanied by the relaxation caused by high concentrations of substance P (SP) and bradykinin (BK). 3. To examine the role played by nitric oxide and prostacyclin in the endothelium-dependent relaxations and hyperpolarizations caused by SP and BK on pig coronary arterial strips, the production of NO was inhibited by NG-nitro-L-arginine (L-NNA) and the production of prostacyclin was inhibited by indomethacin, while monitoring smooth muscle membrane potential and isometric tension. 4. Indomethacin had no effect on resting isometric tension nor on SP and BK relaxations of strips precontracted by prostaglandin F2 alpha. 5. L-NNA contracted arterial strips with intact endothelium, without changing the membrane potential of smooth muscles. 6. The inhibitor shifted to the right the concentration-response curve of kinins by 0.2 nM SP and 20 nM BK. It inhibited the maximal relaxations and hyperpolarizations by 30%. 7. The results show that, in pig coronary arteries, EDRF (NO) mainly controls the basal tension, whereas other factor(s) play(s) an important role in hyperpolarizations and relaxations caused by the kinins.


Ca(2+)-activated and voltage-gated K+ currents in smooth muscle cells isolated from human mesenteric arteries.

1. Smooth muscle cells were enzymatically isolated from arteries dissected from mesenteric fat removed from patients undergoing routine surgery. The whole-cell patch clamp technique was used to characterize the potassium (K+) currents and passive electrical properties of these cells, using high-K(+)-containing pipette solutions with either 0.2 mM EGTA or 10 mM EGTA and 10 mM BAPTA. 2. Cell capacitance, which is proportional to membrane surface area, was normally distributed around a value of 46 pF, and independent of artery size between 0.4 and 3.6 mm. The mean membrane potential measured under current clamp was -44.1 +/- 1.9 mV (n = 52). 3. Cells dialysed with 0.2 mM EGTA in order to weakly buffer intracellular Ca2+ demonstrated a noisy outward current with an apparent threshold near -30 mV, upon which were superimposed spontaneous transient outward currents (STOCs). In the presence, but not the absence, of extracellular Ca2+, this current was potentiated if the holding potential was depolarized into the voltage range between -40 and +50 mV. This potentiation had a bell-shaped potential dependency which reflected the activation of voltage-gated Ca2+ channels in these cells. 4. The noisy current was blocked by externally applied tetraethylammonium (the dissociation constant, Kd = 0.85 mM), as were STOCs. This current was also reduced by about 40% by 8 nM charybdotoxin, and was transiently potentiated by 10 mM caffeine. The characteristics of this current therefore suggested that it was carried by large-conductance Ca(2+)-activated K+ channels. 5. Dialysis of human mesenteric arterial cells with 10 mM EGTA and 10 mM BATPA was not able to completely suppress the Ca(2+)-activated current, and reduced by approximately 50% the amplitude of the outward current recorded at positive potentials. 6. Depolarization of strongly Ca(2+)-buffered cells in the presence of 30 mM TEA to block Ca(2+)-activated K+ channels revealed a residual outward current which had both transient and sustained components. These were blocked by 4-aminopyridine (4-AP) with a similar efficiency (Kd was 1.04 and 1.16 mM at +60 mV for transient and sustained current, respectively), but the voltage ranges over which they inactivated, and their rates of recovery from inactivation, were significantly different. 7. The transient and sustained currents had different sensitivities to external Ca2+ and Cd2+ ions. Ca2+ (5 mM) significantly reduced the amplitude and shifted the voltage dependency of inactivation of the transient but not the sustained component of the outward current. Cd2+ (0.2 mM) reduced the transient current by about 30% without affecting the sustained component amplitude.(ABSTRACT TRUNCATED AT 400 WORDS)


[Serum lipoprotein (A) in patients with myocardial infarction and aortoarteritis].

To elucidate the physiological role of lipoprotein (a), a new independent atherosclerosis risk factor, the levels of lipoprotein (a) and 5 proteins of an acute phase were measured in acute (myocardial infarction, n = 21) and chronic (non-specific aortoarteritis, n = 15) tissue lesions. Blood was taken for analysis from patients with myocardial infarction when they were admitted to hospital for anginal attacks and then 4 times during a month and from patients with aortoarteritis at the stage of exacerbation 1 month before and after therapy. The findings indicate that in acute and chronic tissue lesions, there is a change in blood lipoprotein (a) levels, which is largely similar to that in the levels of acute phase proteins. One cause of such changes may be an increase in the synthesis of apolipoprotein (a) that is identical to that of other glycoproteins--acute phase proteins--in the reaction of the acute phase and then in the regeneration of damaged tissues.


A novel large-conductance Ca(2+)-activated potassium channel and current in nerve terminals of the rat neurohypophysis.

1. Nerve terminals of the rat posterior pituitary were acutely dissociated and identified using a combination of morphological and immunohistochemical techniques. Terminal membrane currents were studied using the 'whole-cell' patch clamp technique and channels were studied using inside-out and outside-out patches. 2. In physiological solutions, but with 7 mM 4-aminopyridine (4-AP), depolarizing voltage clamp steps from different holding potentials (-90 or -50 mV) elicited a fast, inward current followed by a slow, sustained, outward current. This outward current did not appear to show any steady-state inactivation. 3. The threshold for activation of the outward current was -30 mV and the current-voltage relation was 'bell-shaped'. The amplitude increased with increasingly depolarized potential steps. The outward current reversal potential was measured using tail current analysis and was consistent with that of a potassium current. 4. The sustained potassium current was determined to be dependent on the concentration of intracellular calcium. Extracellular Cd2+ (80 microM), a calcium channel blocker, also reversibly abolished the outward current. 5. The current was delayed in onset and was sustained over the length of a 150 ms-duration depolarizing pulse. The outward current reached a peak plateau and then decayed slowly. The decay was fitted by a single exponential with a time constant of 9.0 +/- 2.2 s. The decay constants did not show a dependence on voltage but rather on intracellular Ca2+. The time course of recovery from this decay was complex with full recovery taking > 190 s. 6. 4-AP (7 mM), dendrotoxin (100 nM), apamin (40-80 nM), and charybdotoxin (10-100 nM) had no effect on the sustained outward current. In contrast Ba2+ (200 microM) and tetraethylammonium inhibited the current, the latter in a dose-dependent manner (apparent concentration giving 50% of maximal inhibition (IC50) = 0.51 mM). 7. The neurohypophysial terminal outward current recorded here corresponds most closely to a Ca(2+)-activated K+ current (IK(Ca)) and not to a delayed rectifier or IA-like current. It also has properties different from that of the Ca(2+)-dependent outward current described in the magnocellular neuronal cell bodies of the hypothalamus. 8. A large conductance channel is often observed in isolated rat neurohypophysial nerve terminals. The channel had a unit conductance of 231 pS in symmetrical 150 mM K+.(ABSTRACT TRUNCATED AT 400 WORDS)


Reversal of hypotension induced by Vibrio vulnificus lipopolysaccharide in the rat by inhibition of nitric oxide synthase.

Intravenous infusion of Vibrio vulnificus lipopolysaccharide (LPS) (1 mg/kg body wt) in rats caused a dramatic drop in mean arterial pressure within 10 min and a further decline in mean arterial pressure and heart rate which lead to death between 25 and 70 min. Rats treated with LPS followed 10 min later by the intravenous infusion of NG-monomethyl-L-arginine (L-NMMA, 20 mg/kg body wt) showed an initial drop in mean arterial pressure owing to the LPS infusion, followed by a transient rise in mean arterial pressure which lasted for approximately 40 min after the infusion of L-NMMA. The pressure values then remained level for at least 150 min post-LPS infusion. Control rats treated with equivalent volumes of saline infusion showed stable values of mean arterial pressure and heart rate. Additional control rats receiving L-NMMA alone showed the transient rise in mean arterial pressure, followed by a return to the baseline values. The results indicate that the symptoms of endotoxic shock resulting from V. vulnificus LPS may result in part from the stimulation of the activity of nitric oxide synthase. Inhibition of nitric oxide synthase by L-NMMA is a possible treatment for toxic shock induced by V. vulnificus.


The prevalence of the indirect hemagglutination test for melioidosis in children in an endemic area.

The indirect hemaglutination test for melioidosis was studied in 295 children who live in the northeastern part of Thailand. Sixty-seven children (22.7%) were healthy children who came to the well baby clinic. Two hundred and twenty-eight children (77.3%) came to the hospital because of some illnesses other than melioidosis. Eighty-three percent of the children had an IHA titer of at least 1:10 or greater. Twenty-two percent had an IHA titer of 1:80 or greater. The prevalence of positive IHA titer and the mean titer were higher in the older age group. The age of children should be considered when interpreting IHA titer for milioidosis.


Experimental omento-myelo-synangiosis.

In 18 cats the omentum was mobilized, transposed, and placed directly upon the pia mater of the spinal cord. Within 2 months, vascular anastomoses developed at the omento-spinal cord interface and, as demonstrated by India ink perfusion, communicated widely with the cord's intramedullary vessels. Light and scanning electron microscopy (SEM) of the interface revealed the development of a "fibrous coat" of Goldsmith in continuity with the edges of the dura mater. It consisted mainly of a matrix of collagen fiber and fibroblasts, well vascularized with sinusoids, capillaries, arterioles and venules. The sinusoids, as identified by SEM, were present in both the fibrous coat and adjacent omentum, were lined with attenuated endothelium, and had multiple communications. The possible significance of the sinusoidal formation is discussed in this report.


Study of bovine herpesvirus type 1 strains with monoclonal antibodies.

Fourteen strains of bovine herpesvirus type 1 (BHV-1, IBRV) representing all three groups of BHV-1 (BHV-1.1, BHV-1.2, BHV-1.3) were studied by ELISA using 106 monoclonal antibodies (Mabs) produced against BHV-1. On the basis of the ELISA, the Mabs could be divided into three groups. The first group (40 Mabs, 38%) reacted with all strains, the second group (43 Mabs, 41%) with the respiratory and genital strains (BHV-1.1 and BHV-1.2) while the third group (23 Mabs, 22%) only with the respiratory strains. Only 5 out of the antibodies neutralized respiratory and genital strains, and none of them neutralized the encephalitogenic strains (1.3). Three Mabs selected from each of the 3 groups, and the above five neutralizing strains were studied by Western blot. Antibodies of groups 1 and 3, and two neutralizing antibodies bound to a 90k protein (gpIII), whereas members of group 2 and 3 neutralizing antibodies reacted with a 74k and a 130k protein (both gpl). The results indicate that reactivity with monoclonal antibodies is as suitable for the classification of BHV-1 strains as is restriction endonuclease (RE) analysis but it cannot distinguish between subgroups within the groups.


[Pneumocystis carinii pneumonia: laboratory diagnosis].

We worked with 51 samples, 7 bronchoalveolar lavages (BAL) and 44 sputa (S) of 31 AIDS patients with clinical and radiographic symptoms compatible with Pneumocystis pneumonia. With the aim of finding a specific sensitive methodology for the diagnosis of Pneumocystis carinii, we evaluated 4 coloration techniques (silver methenamine, its modification without gold chloride, toluidine blue and Giemsa). 35% of the patients studied were positive. P. carinii were observed in 18% of the 44 sputa. We observed that the analysis of a single sputum sample (S) has a very low sensitivity and that the processing of two or more samples is necessary since only one of the 14 patients who had sent a single sample was found P. carinii positive, while in the remaining ten who had sent more than one (S) sample, the microorganism was detected in 50%. 4 of the 7 BAL were positive. 4 BAL were preceded by the analysis of an (S) sample: in two cases the results were negative while BAL allowed us to make the diagnosis, thus demonstrating its greater efficacy. To enhance sensitivity each sample was centrifuged until exhaustion and 10 slides were prepared for coloration with the final sediment. The four techniques employed were specific and all the Pneumocystis pneumonia patients responded to the treatment. Silver methenamine, its modification without gold chloride, and toluidine blue were very sensitive, in contrast to of Giemsa. The stain to be chosen is either silver methenamine, or its modification, because both achieve the best contrast, allowing optimum P. carinii identification. We suggest the implementation of some of these techniques in laboratory routine.


[Cardiovascular complications of interferon].

Interferon, a fairly recent drug, is used in some cancers and other diseases. The adverse effects include cardiotoxicity which was recognized from phase-I trials. Some of these complications are very common and not serious at the dose levels administered: transient hypotension at the beginning of the treatment, atrial extrasystole. Other effects are less common and not dangerous: low-level conduction impairment or reversible hypertension as the authors recalled. There are also a much more serious forms of toxicity which may be life-threatening, one of which seems to be dose-dependent and consists of the onset of cardiomyopathy, which is usually reversible when treatment is stopped, the other is uncontrollable and usually occurs in high-risk cardiac patients from the first few injections and results in sudden death induced by acute coronary artery failure and/or serious ventricular arrhythmia. The physiopathology of this cardiotoxicity remains unknown, but as it is known to exist, rigorous cardiological monitoring of all patients receiving this treatment is necessary.


Investigation of the solid- and solution-phase binding reactivities of continuous epitopes recognized by polyclonal guinea-pig anti-recombinant bovine growth hormone antisera.

We have used the technique of multiple pin peptide synthesis to identify three major continuous epitopes in the recombinant bovine (rb) GH molecule. We have synthesized these peptides, residues 24-40, 139-152 and 179-189, as N-terminally acetylated, C-terminal amides and confirmed their reactivity in a standard solid-phase ELISA. Subsequently, for epitope 139-152, we have synthesized a peptide affinity column and used this to isolate antibodies with this epitope specificity from whole antiserum. In addition, we demonstrate that under native conditions in a liquid phase RIA, these antibodies will precipitate [125I]rbGH. Further, peptide 139-152 itself also cross-reacts in an rbGH RIA inhibiting binding by up to 20%. Our data suggest that during the immune response to rbGH in guinea-pigs a substantial part of the B-cell response is directed to the 139-152 region and that this part of the protein is a native epitope.


Role of autocrine and paracrine factors in thyroid follicle growth.

Basic fibroblast growth factor (bFGF), insulin like growth factor I and II and transforming growth factor beta (TGF-beta) are assumed to be of importance in the paracrine and autocrine regulation of thyroid growth and function. Using in vitro cultures of isolated intact porcine thyroid follicles, we here present data that support a possible autocrine action of bFGF on proliferation, a possible explanation for the observed potentiation of EGF-stimulated growth by IGF-I, and results on the release and regulation of release of TGF-beta. For growth experiments, thyroid follicles (2 x 10(5) cells) were incubated for 6 days followed by cell counting. For the analysis of EGF binding sites, follicles were preincubated with and without IGF-I (10 ng/mL) for 48 h at 37 degrees C, incubated with 125I-EGF (5 nCi/well) and unlabeled EGF (0.1-500 ng/mL) for 24 h at 4 degrees C (2 x 10(5) cells/well); binding characteristics were calculated from Scatchard analysis. The TGF-beta bioactivity in untreated and acid treated media conditioned with thyroid follicles for 3 days (2 x 10(7) cells) was analyzed with a bioassay using mink lung epithelial cells. Basic FGF (0.1-1 ng/mL) dose-dependently stimulated the proliferation of thyroid follicles up to 135.0 +/- 6.1%; this effect was additive with IGF-I (10 ng/mL) but not with EGF (2 ng/mL). The IGF-I (10 ng/mL) just moderately increased proliferation (128.3 +/- 16%), but potentiated EGF (1 ng/mL)-stimulated growth (from 183.0 +/- 11.0% to 314.0 +/- 3.0%).(ABSTRACT TRUNCATED AT 250 WORDS)


Effects of pineal-derived indolic compounds and of certain neuropeptides on the growth processes in the thyroid gland.

The aim of the present study was to examine the effects of melatonin (Mel) and of pinealectomy (PX) [in a long-term experiment in vivo - 10 weeks], as well as of Mel and N-acetylserotonin (NAc-5HT) [in experiments ex vivo in vitro and in vitro], on the rat thyroid growth processes. Additionally, the incubations in vitro of rat thyroid lobes with 3H-thymidine, in the presence of TSH, vasoactive intestinal polypeptide (VIP), VIP-antagonist ([4Cl-D-Ph6, Leu17]VIP), somatostatin (SS), all the substances used separately or jointly in combinations, were performed. It was shown that: (a) Mel--administered in late afternoon injections--decreased, while PX increased examined indices of thyroid growth in vivo, (b) Mel--administered in s.c. implanted pellets--reversed the inhibitory effect of Mel injections, (c) in experiments ex vivo in vitro and in vitro, the inhibitory effect of Mel revealed only for the lowest applied dose/concentration of the hormone, (d) NAc-5HT showed no effect, (e) VIP decreased 3H-thymidine incorporation into DNA of thyroid lobes in vitro and enhanced the inhibitory effect of SS on the process in question, (f) VIP-antagonist failed to reverse the inhibitory action of VIP on the thyroid growth.


Selenium supply regulates thyroid function, thyroid hormone synthesis and metabolism by altering the expression of the selenoenzymes Type I 5'-deiodinase and glutathione peroxidase.

Selenium supply appears to be insufficient in several regions of Europe as indicated by analysis of plasma and tissue selenium content and measurements of selenium dependent glutathione peroxidases (GPx). Selenium deficiency alters both thyroid hormone synthesis and tissue specific activation by 5'deiodinase isoenzymes. Human and rat Type I 5'deiodinase have been identified as selenoprotein containing selenocysteine in its active site. Regulation of selenium dependent expression of Type I 5'deiodinase occurs at the transcriptional and posttranscriptional level and a cellular hierarchy of selenite incorporation was found for Type I 5'deiodinase over GPx in LLC-PK1 kidney cells. The role of other newly discovered selenoproteins for thyroid hormone synthesis, activation, and action has to be investigated.


Iodine and goiter involution.

Iodine administration, although efficient in goiter treatment or prevention, is also responsible for adverse effects such as cell necrosis or thyroiditis. These two effects were reproduced in iodide-treated goitrous mice. Morphological observations strongly suggest that thyroid cell death results from an excessive production of free radicals, which initiates lipid peroxidation. This hypothesis is strengthened by the facts that the thyroidal concentration of malonic dialdehyde, a stable product of lipid peroxidation, is increased, and that necrosis is partially prevented by free radicals scavengers. Epithelial necrosis is associated to an inflammatory reaction. The infiltrate is mainly made of cells expressing class II molecules of major histocompatibility complex (macrophages and dendritic cells), but also of T lymphocytes. However, this inflammation, which varies among mouse strains, is transient and it is not amplified or maintained by administration of cytokines, IFN gamma or TNF alpha, known to induce class II expression on thyrocytes.


Further studies on the antigoitrogenic action of iodoarachidonates.

Previous studies have shown that iodoarachidonates (IAs) prevent goiter production in rats. In the present studies we show that both IL-d and IL-w (IAs bearing the iodine atom at the positions 6 and 14, respectively), cause a significant involution of preformed goiter. This effect was evident when IAs were administered either orally or via i.p., although the first one required larger doses to obtain the same degree of inhibition. No changes were observed in serum protein, urea, cholesterol, cholinesterase, T3 or T4. In vitro studies with FRTL-5 cells showed that both IAs inhibit iodide and alpha-AIB uptake, as well as ATPase activity.


The role of iodine and thyroid cell growth.

The autoregulatory effects of iodide on thyroid growth and function are discussed to be mediated by iodinated derivatives of essential fatty acids (EFA), esp. iodolactones. We now reevaluated the effect of iodide on proliferation of isolated porcine thyroid follicles by determination of cell counts and investigated the effects of pretreatment of the follicles with arachidonic acid (AA, C 20:4 n6) in comparison to docosahexaenoic acid (DHA, C22:6 n3). Growth experiments were performed in multi-well culture plates and cell counts were determined after 6 d of incubation. EGF (5 ng/ml) significantly stimulated thyroid cell proliferation (151 +/- 6%; Mean +/- SD vs. basal control, 100 +/- 8%). 2.5 microM of iodide, added 24 h before EGF, had a weak stimulatory effect (168 +/- 9%) whereas higher concentrations of iodide (5-80 microM) exerted significantly dose-dependent inhibitory effects (117 +/- 1% at 80 microM of KI) which could be abolished with 500 microM of methimazole (155 +/- 11% at 80 microM of KI). Isolated porcine follicles showed a rapid uptake of EFA (25 microM) measured by specific tracer activity in the ethanol/acetic acid extracts of follicles (1.60 +/- 0.48 mumol EFA/ml follicle/24 h). Treatment with DHA (100 and 300 microM) significantly enhanced the inhibitory effect of 10 microM of iodide on thyroid follicle proliferation (84 +/- 2% and 45 +/- 4%) in contrast to follicles pretreated with AA (100 +/- 8% and 60 +/- 8%). These results demonstrate the biphasic effect of iodide on thyroid growth which can be abolished by inhibition of iodide organification with methimazole.(ABSTRACT TRUNCATED AT 250 WORDS)


Treatment of iodine deficiency goiter with iodine, levothyroxine or a combination of both.

During recent years several studies have been published comparing different ways of pharmacological treatment of a goiter due to iodine deficiency. These studies usually were performed with 300 to 500 micrograms of iodine, 100 to 150 micrograms levothyroxine, or a combination of in most cases 100 micrograms levothyroxine and 100 micrograms iodine. The largest data have been accumulated in 166 patients with in most cases diffuse goiter. Group A (n = 61) received 150 micrograms levothyroxine per day, group B (n = 50) 400 micrograms iodine per day and group C (n = 55) a combination of 75 micrograms levothyroxine and 200 micrograms iodine per day. During the eight months of therapy, in all three groups a significant and comparable mean decrease in goiter size was documented (-32.1% in group A, -37.3% in group B and -38.7% in group C [n.s. between the three groups]). Striking differences between the three groups are evident in the changes of basal and thyrotropin releasing hormone (TRH) stimulated thyrotropin (TSH). In group A, after eight months a sharp and significant decrease of TSH occurred (from 1.2 mU/l to 0.4 mU/l; mean; p < 0.05), while in group B TSH showed only a minor decrease (from 1.3 mU/l to 0.9 mU/l) and remained significantly higher compared to both, group A and C (p < 0.01). Similar changes were documented when the TSH after TRH administration was calculated. It is concluded, that all three therapeutic approaches are effective for goiter reduction.(ABSTRACT TRUNCATED AT 250 WORDS)


Determinants of outcome in sporadic nontoxic goiter.

The author points out that natural history of sporadic nontoxic goiter (SNG) is characterized by increasing thyroid volume, nodularity and autonomy of function leading to hyperthyroidism in 13-16% of the patients after a medium follow-up of circa 12 years. Successively, the author reports and discusses the effectiveness and the outcome of the therapeutical modalities used for SNG: thyroidectomy, thyroid hormones and radioactive iodine.


Interactions between the immune system and the thyroid. Regulatory networks in health and disease.

Hashimoto's thyroiditis, primary myxedema and Graves' disease are thyroid disease that are due to autoimmune reactions towards thyroidal antigens such as thyroid peroxidase (TPO), thyroglobulin (Tg) and the TSH receptor. Thyrocyte destruction in Hashimoto's thyroiditis and primary myxedema is caused by TPO- and Tg-specific lymphocytes and autoantibodies, thyrocyte stimulation in Graves' disease is caused by antibodies stimulating the TSH receptor, thyroid atrophy in primary myxedema is caused by antibodies blocking the TSH receptor, or a yet unknown thyroid growth receptor. The above listed thyroid autoimmune diseases are familial (genetically determined), and due to defects in the immunoregulatory mechanisms that should normally control excessive thyroid autoimmune reactivity. This control towards thyroidal antigens (tolerance) can be broken by professional antigen presenting cells, such as the dendritic cells. It is now known that thyroid autoimmune diseases are indeed initiated by dendritic cells: dendritic cells are present in low number in normal thyroids, but accumulate very early in thyroids that are later affected by thyroid autoimmune disease. Dendritic cells are also present in the normal anterior pituitary and in this gland they are known as the (folliculo) stellate cells, the regulators of growth and function of the surrounding pituitary-endocrine cells. It is discussed whether the influx and clustering of dendritic cells in the thyroid observed during early autoimmune thyroid disease is meant for the regulation of growth and function of the thyrocytes thus linking a putative early endocrine disturbance to the initiation of thyroid autoimmune disease.


Use of FRTL-5 for the study of thyroid antibodies involved in goitrogenesis.

Some authors have suggested a role of autoimmunity in the pathogenesis of iodine deficiency disorders (IDD). For this purpose we have searched for thyroid adenylate cyclase stimulating antibody (TSAb) and thyroid growth stimulating antibody (TGSAb) in patients with endemic goiter (EG) and endemic cretinism (EC). Immunoglobulins G preparations (IgGs) were tested in FRTL-5 cells. TSAb were calculated as percent of cAMP increase over basal production and TGSAb were expressed as percent of increase of 3H-thymidine incorporation and DNA content in FRTL-5 cells. Our results show that IgGs from goitrous patients were devoid of TSAb and TGSAb activities, while in the same conditions IgGs from patients with Graves' disease had the ability to stimulate cAMP production and 3H-thymidine incorporation in FRTL-5 cells. These data argue against a direct role of TSAb and TGSAb in the pathogenesis of IDD.


Involvement of the immune system in iodine deficient goiter.

Iodine deficient goiters were studied by immunohistochemistry and showed extensive presence and typical arrangement of dendritic cells, known to have excellent antigen presenting capacity. These cells were positive for all MHC-class II epitopes and for ICAM-1. Epithelial follicle lining cells were also seen to be class II positive but lacked ICAM-1. Thyroglobulin seemed not to be iodinated at the C-terminal hormogenic site, as shown by reactions with monoclonal antibodies. Iodine therapy, as well as thyroxine therapy were effective in reducing thyroid size. Both forms of therapy were found to decrease the pretreatment levels of circulating thyroid growth stimulating immunoglobulins (TGI).


Gi alpha-1 expression in the human thyroid is regulated by TSH: loss of regulation in thyroid autonomous adenoma.

The molecular mechanisms underlying the development of endocrine active thyroid tumors are poorly understood. These tumors produce excess thyroid hormone, which then suppresses TSH (thyroid stimulating hormone) production. In the present report, we show that the expression of Gi alpha-1 is under control of TSH in the normal human thyroid. In contrast Gi alpha-1 escapes TSH control in autonomous adenoma and thus is constitutively expressed. Since receptor-mediated activation of Gi controlled pathways is known to elicit a proliferative response in several cell types, we propose that in thyroid adenomas the unregulated constitutive expression of Gi alpha-1 is causally related to the autonomous growth.


The hare parotid gland: ultrastructure and histochemistry of acinar and ductal cells.

The parotid gland of the hare Lepus europaeus was studied by electron microscopy. The parenchyma consists of secretory areas (acini) and ductal segments (intercalated, striated and excretory ducts). Acinar cells showed a marked heterogeneity due to the presence of secretory granules characterized by different size, electron-density and degree of membrane merging in addition to the occurrence of flattened and dilated rough endoplasmic reticulum. Acinar cells had variable affinity for PAS and PA-TCH-SP staining but were HID- and LIT-TCH-SP negative. Intercalated duct cells lacked secretory granules. Striated ducts were lined by four cell types namely light cells, dark cells, vacuolated cells, and typical striated cells. Excretory ducts showed some scalloping on the apical region of the light cells.


[Effects of combination chemotherapy of ovarian cancer in vitro].

To investigate the effects of combination chemotherapy on ovarian cancer in vitro, we observed the sensitivity of tumors to different drugs by incorporation assay of tritium thymidine (3H-TdR). The results revealed that the drugs and their sensitive degrees varied from tumor to tumor. Whether the combination of the drugs is synergism or sometimes only enhances toxicity depends on the difference of individuals. We suggest that in vitro anti-cancer drug sensitivity test could be applied in selection or determination of chemotherapy protocol.


Human immunodeficiency virus in gestational trophoblastic neoplasias--is it a poor prognostic risk factor.

Three cases of HIV infection with choriocarcinoma are presented. One case had prolonged chemotherapy without remission, the second had remission only after combining hysterectomy with chemotherapy and the third who had extensively metastatic disease in the presence of other low risk factors are reported. HIV infection may predispose patients to extensive metastatic choriocarcinoma and influence the course of treatment. We propose that HIV infection be considered a poor prognostic risk for gestation trophoblastic neoplasias.


[The effect of substance P on histamine-stimulated gastric juice release and bile formation].

The substance P was shown to be able to increase the level of histamine-induced stomach secretion and the debit of free hydrochloric acid in dogs. This peptide increases the volume velocity of the choleresis and the content of bile acids and protein in the bile. The substance P alters the range of the bile acids as well.


[Comparative studies on LH/CG receptor from rat testes and hCG-binding protein from Pseudomonas maltophilia by antibodies against the hCG-binding protein].

An indirect immunoprecipitation experiment was developed to characterize the antigenic determinants of the hCG-binding protein and the rat testicular LH/CG receptor. The hCG-binding protein preincubated with 125I iodo hCG was immunologically precipitated by the antibodies against the hCG-binding protein in the presence of goat anti-rabbit IgG. Similarly, the LH/CG receptor solubilized with 1% triton X-100 from the rat testes was also immunoprecipitated by the antibodies. Furthermore, the antibodies did not inhibit the binding of 125I iodo hCG to the hCG-binding protein or to the solubilized rat testicular LH/CG receptor. From these results it can be concluded that there exists the sharing of antigenic determinants between the hCG-binding protein from Pseudomonas maltophilia and the LH/CG receptor from the rat testes and that the antibodies specifically recognize the antigenic determinants outside the hCG-binding sites on the rat testicular LH/CG receptor. Therefore, the antibodies against the hCG-binding protein are of potential applications to studies on structure and function of the mammalian LH/CG receptors.


[A randomized control trial of isovolemic hemodilution therapy in cor pulmonale].

The hemorheologic properties are obviously abnormal in patients with cor pulmonale (CP), presenting as high hematocrit (HCT), hyperviscosity and hyper-viscoelasticity. Twenty patients suffering from CP with polycythemia were randomized into either an isovolemic hemodilution (IHD) group or control group. In the IHD group, 10 patients were treated by routine therapy combined with IHD. The other 10 patients in the control group were treated by routine therapy only. The results demonstrated that after treatment in IHD group, all viscoelasticity property parameters except eta p were decreased significantly. Oxygen delivery, oxygen transport capacity, and stroke volume and cardiac output were increased. Mean pulmonary artery pressure and total pulmonary resistance were decreased. However, in the control group only HCT and eta 0.512 were decreased after treatment. The viscosity and eta' of the whole blood in IHD group were decreased more markedly than those in the control group. IHD was well tolerated in most of the patients. There were no complications or side effects. Thus, IHD is a safe, effective and acceptable therapy, and it may play a role in the treatment of cor pulmonale.


Utilization of the Boyden chamber to further characterize in vitro migration and invasion of benign and malignant human prostatic epithelial cells.

In the present study, we assessed various tissue culture conditions on the motility and invasiveness of benign and malignant prostatic epithelial cells using the Boyden chamber system. In DME and RPMI-1640 media, benign human prostatic epithelial cells were unable to penetrate through a basement-membrane-coated filter. However, the prostatic carcinoma cell lines, PC3 and DU145, showed a significant frequency of invasion under those conditions. When cultured in the low-calcium WAJC 404 medium, benign prostatic epithelial cells efficiently penetrated through the basement-membrane-coated filter. PC3 and DU145 cells showed an attenuated capability to invade when cultured in WAJC 404 medium. The effect of extracellular calcium on the behavior of these cells in the Boyden chamber was further evaluated by gradual addition of calcium chloride in WAJC 404 medium. These studies showed that the number of benign prostatic epithelial cells penetrating through the filter decreased as the calcium ion concentration increased. Conversely, the number of PC3 cells invading through the filter increased as calcium ion concentration was increased to 0.4 mM and decreased at higher calcium ion concentrations. These results suggest that the extracellular calcium concentration is one of the factors which may affect cell behavior in the Boyden chamber system.


Effect of injury on pulpal levels of immunoreactive substance P and immunoreactive calcitonin gene-related peptide.

Neuropeptides such as calcitonin gene-related peptide (CGRP) and substance P are present in dental pulp in relatively high concentrations. Previous studies have demonstrated that the staining density of immunoreactive CGRP (iCGRP) changes in dental pulp after tissue injury. This study evaluated injury-related changes in levels of both immunoreactive CGRP (iCGRP) and immunoreactive substance P (iSP) in dental pulp using radioimmunoassays. After pulpal exposure, iSP levels decreased to about 10% of baseline values, while iCGRP levels decreased to about 45% of baseline measures. After dentin exposure with acid etch, iSP levels decreased to about 10 to 20% of baseline measures, while iCGRP levels decreased to 60% of baseline values. For both forms of injury, iSP decreased to a greater extent than did iCGRP levels. Collectively, these findings indicate that pulpal neuropeptides undergo dynamic, injury-specific, and peptide-specific responses following trauma to dental pulp.


An in vitro method to evaluate regulation of neuropeptide release from dental pulp.

Although pulpal neuropeptides such as calcitonin gene-related peptide and substance P may mediate neurogenic inflammation, little is known about the regulation of neuropeptide release from dental pulp. This article describes an in vitro method for superfusing dental pulp which permits the study of mechanisms regulating the release of immunoreactive CGRP (iCGRP). Tissue extracts from bovine dental pulp dilute in parallel to authentic calcitonin gene-related peptide and substance P peptide standards when assayed by radioimmunoassay. Pulpal levels of iCGRP were 17-fold greater than levels of immunoreactive substance P. Administration of a potassium pulse evoked a significant release of iCGRP from dental pulp (155 +/- 21 fmol/g/9 min) as compared with iCGRP spontaneously released from concurrent control chambers (18 +/- 11 fmol/g/9 min). The in vitro superfusion of pulp tissue may serve as a useful method for identifying peripherally acting drugs which modulate nociceptor secretory activity and for determining their mechanisms of action.


[The extracellular ionic environment and bone metabolism. In vitro study of the role of chloride].

Controversies surround the view that the blood-bone ionic equilibrium is controlled by ionic changes at a so called "bone membrane" that display a pump-leak mechanism able to maintain a concentration gradient for chloride in addition to that for calcium, sodium and potassium. On the other hand it is known that a chloride-bicarbonate transport is active at bone cells membrane to control cytosolic pH. We tested the hypothesis that a modification of the chloride concentration of the culture medium, as well as a block of the chloride-bicarbonate transport by an anionic channels blocker, in an experimental model that keeps intact the bone membrane, may influence the bone cells activity and that the influence may be quantitatively different to that previously observed in isolated bone cells. The experimental model is the intact fetal rat limb bone, cultured for 24 hours in a simplified medium at different chloride concentrations obtained by substituting chloride with isethionate. Alkaline phosphatase activity released in the medium was measured as a marker of osteoblasts activity. By progressively reducing the chloride concentration from 117 mM to 29 mM, alkaline phosphatase activity was found unchanged. By dramatically reducing the medium chloride concentration to the range 14-0 mM, alkaline phosphatase activity was significantly (p = 0.0002) inhibited by 31.58%.(ABSTRACT TRUNCATED AT 250 WORDS)


[Development of a system for cloning a DNA fragment, containing the determinant of resistance to actinomycin for Streptomyces chrysomallus No.2--a producer of actinomycin C].

To study the modes of actinomycin biosynthesis and the mechanism responsible for resistance to the antibiotic producing S. chrysomallus No. 2, the authors undertook an examination and studies into the cloning system for gene(s) of resistance to actinomycin from a S. chrysomallus No. 2 actinomycin C producer and the cloning of a S. chrysomallus No. DNA fragment to the actinomycin-sensitive Streptomyces Sp. 26-115 H-I on the vector plasmid pIJ702. The cloning gave rise to actinomycin-resistant strains. The character of actinomycin resistance is inheritable in a steady fashion.