A high-throughput assay for screening the abilities of per- and polyfluoroalkyl substances in inducing plasma kallikrein-like activity.

The plasma consists of multiple functional serine zymogens, such as plasma kallikrein-kinin system (KKS), which are vulnerable to exogenous chemical exposure, and may closely relate to the deleterious effects. Testing whether the anthropogenic chemicals could increase the kallikrein-like activity in plasma or not would be of great help to understand their potentials in triggering the cascade activation of the plasma zymogens and explain the corresponding hematotoxicity. In this study, a novel high-throughput ex vivo assay was established to screen the abilities of emerging chemicals like per- and polyfluoroalkyl substances (PFASs) in inducing kallikrein-like activities on basis of using rat plasma as the protease zymogen source. Upon the optimization of the conditions in the test system, the assay gave sensitive fluorescent response to the stimulation of the positive control, dextran sulfate, and the dose-response showed a typical S-shaped curve with EC50 of 0.24 mg/L. The intra-plate and inter-plate relative standard deviations (RSDs) were less than 10% in the quantitative range of dextran sulfate, indicating a good reliability and repeatability of this newly-established assay. Using this method, several alternatives or congeners of perfluorooctanesulfonic acid (PFOS) and perfluorooctanoic acid (PFOA), including 6:2 chlorinated polyfluoroalkyl ether sulfonate (6:2 Cl-PFESA), Ag-PFOA, K-PFOA, Na-PFOA and ammonium pentadecafluorooctanoate (APFO), were further screened, and their capabilities in inducing kallikrein-like activities were identified. The ex vivo assay newly-developed in the present study would be promising in high-throughput screening of the hematological effects of emerging chemicals of concern.

Effect of Repeated Administration of Cefquinome on Biochemical and Hematological Parameters in Buffalo Calves.

AIM: Cefquinome, a fourth generation of cephalosporins have been developed for use in animals. Similar to other species, it may also have some adverse reactions in buffalo calves at therapeutic dosage. In the present study, effect of repeated administration of cefquinome on biochemical and hematological parameters was studied in buffalo calves. MATERIALS AND METHODS: Animals were divided into two groups having three animals in each group. Group 1 was kept as control and animals of Group 2 were given cefquinome at dose rate of 2 mg.kg(-1) body weight by intramuscular route for continuously 7 days. Blood samples were collected daily and 3 days post treatment. RESULTS: The values of aspartate aminotransferase (AST), alanine aminotransferase (ALT), gamma-glutamyl transpeptidase (GGTP), and alkaline phosphatase (ALKP) in control animals were in the range of 127.7-148.3 IU/L,49.0-55.6 IU/L, 14.0-17.3 IU/L, and 111.0-134.3 IU/L, respectively. The repeated administration of cefquinome did not influence the plasma activities of AST, ALT, GGTP, and ALKP in treated animals. The level of blood urea nitrogen (BUN) and creatinine before treatment was 14.3 ± 0.88 mg/dl and1.70 ± 0.04 mg/dl, which significantly increased on 3(rd) day (21.0 ± 1.53 mg/dl) and 2(nd) day (2.33 ± 0.07 mg/dl), respectively. Among hematological parameters, there was significant variation in levels of hemoglobin (Hb), total erythrocyte count (TEC), erythrocyte sedimentation rate (ESR), mean corpuscular volume (MCV), mean corpuscular hemoglobin (MCH), and mean corpuscular hemoglobin concentration (MCHC) in treated animals. No abnormal clinical symptoms were observed in any animal. CONCLUSION: The results revealed that clinically, the therapy of cefquinome may be continued up to 7 days.

Acute exposure to realistic concentrations of Bisphenol-A trigger health damage in fish: Blood parameters, gene expression, oxidative stress.

Despite much information regarding BPA toxicity in fish and other aquatic organisms, data is still misleading as most studies have utilized concentrations several orders of magnitude higher than those typically found in the environment. As an illustration, eight of the ten studies investigating the impact of BPA on the biochemical and hematological parameters of fish have employed concentrations on the order of mg/L. Therefore, the results may not accurately represent the effects observed in the natural environment. Considering the information above, our study aimed to 1) determine whether or not realistic concentrations of BPA might alter the biochemical and blood parameters of Danio rerio and trigger an inflammatory response in the fish liver, brain, gills, and gut and 2) determine which organ could be more affected after exposure to this chemical. Findings pinpoint that realistic concentrations of BPA prompted a substantial increase in antioxidant and oxidant biomarkers in fish, triggering an oxidative stress response in all organs. Likewise, the expression of different genes related to inflammation and apoptosis response was significantly augmented in all organs. Our Pearson correlation shows gene expression was closely associated with the oxidative stress response. Regarding blood parameters, acute exposure to BPA generated biochemical and hematological parameters increased concentration-dependent. Thus, it can be concluded that BPA, at environmentally relevant concentrations, threatens aquatic species, as it prompts polychromasia and liver dysfunction in fish after acute exposure.

Hematological effects of glyphosate in mice revealed by traditional toxicology and transcriptome sequencing.

The herbicide glyphosate is being used worldwide. Hematological toxicity caused by glyphosate exposure has been reported, but the underlying mechanisms remain unclear. In this study, classical toxicology methods and RNA sequencing were performed to explore the molecular mechanisms related to glyphosate hematotoxicity. We found that 500 mg/kg b.w. glyphosate-based herbicide (GBH) significantly decreased leukocyte, neutrophil, lymphocyte and monocyte counts, as well as inhibited colony-forming abilities of CFU-GM, CFU-G and CFU-GEMM. RNA sequencing identified 82 and 48 differentially expressed genes (DEGs) in BM cells after treatment with 250 mg/kg and 500 mg/kg GBH, respectively. Meanwhile, GO and KEGG analyses revealed that the MAPK signaling pathway, hematopoietic cell lineage and cytokine-cytokine receptor interactions were vital pathways involved in GBH-induced toxicity in BM cells. Notably, Nr4a, Fos, Thbs1 and tnfrsf19 contributed to the hematotoxicity of GBH by regulating hematopoietic stem cell functions. In summary, our efforts enhance the understanding of the glyphosate hematotoxic responses and facilitate future studies on its corresponding mechanisms.

Oxidative stress, metallomics and blood toxicity after subacute low-level lead exposure in Wistar rats: Benchmark dose analyses.

Exposure to lead (Pb) is still rising concern worldwide, having in mind that even low-dose exposure can induce various harmful effects. Thus, in-depth knowledge of the targets of Pb toxicity and corresponding mechanisms is essential. In the presented study, the six groups (male Wistar rats, n = 6) received 0.1; 0.5; 1; 3; 7; 15 mg Pb/kg body weight/day for 28 days, each day by oral gavage, while the control group received distilled water only. All animals were sacrificed 24 h after the treatment, and blood was collected for the analysis of hematological, biochemical, oxidative status and essential elements levels. An external and internal dose-response relationship was performed using PROASTweb 70.1 software. The results showed that low doses of Pb affect hematological parameters and lipid profile after 28 days. The possible mechanisms at examined Pb dose levels were a decrease in SOD, O2•- and Cu and an increase in Zn levels. The dose-dependent nature of changes in cholesterol, HDL cholesterol, O2.-, SOD, AOPP in serum and hemoglobin, Fe, Zn, Cu in blood were obtained in this study. The most sensitive parameters that were alerted are Cu blood levels (BMDL5: 1.4 ng/kg b.w./day) and SOD activity (BMDL5: 0.5 µg/kg b.w./day). The presented results provide information that may be useful in further assessing the health risks of low-level Pb exposure.

Evaluation of postoperative pain and toxicological aspects of the use of dipyrone and tramadol in cats.

OBJECTIVES: The aim of this study was to evaluate the effects of dipyrone and tramadol, used for 5 days, on postoperative pain, hematological and biochemical parameters, and oxidative markers on erythrocytes. METHODS: Twenty-eight healthy cats underwent ovariohysterectomy and were randomly allocated to four groups (each n = 7), according to the postoperative treatment administered intravenously: control (saline 1 ml q8h), DIP1 (dipyrone 25 mg/kg q24h), DIP2 (dipyrone 25 mg/kg q12h) and DIP3 (dipyrone 25 mg/kg q8h). All animals received tramadol (2 mg/kg q8h). Pain was assessed by visual analog (VAS), multidimensional UNESP and Glasgow pain scales for cats preoperatively and at 3, 6, 12, 24, 36 and 48 h after extubation. Venous blood was collected daily for 5 days, and on day 10, to perform a complete blood count (CBC) and determine the percentage of Heinz bodies (HBs). Serum biochemistry was evaluated preoperatively and on days 5 and 10; superoxide dismutase (SOD), catalase (CAT), myeloperoxidase (MPO) and lipoperoxidation were evaluated preoperatively and on days 3, 5 and 10. RESULTS: Control cats had higher pain scores than DIP3 cats by UNESP (P = 0.0065), and DIP2 (P = 0.0035) and DIP3 cats (P = 0.0108) by VAS 3 h postoperatively. Rescue analgesia was required by two animals in the control group and one each in the DIP1 and DIP2 groups. There was no difference in SOD or CAT among groups. On day 5, MPO was more active in DIP2 than in DIP3 cats (P = 0.0274). No difference in lipoperoxidation among treatment and control cats was found. CBC remained constant and without statistical difference among groups. Control, DIP2 and DIP3 cats presented a similar percentage of HBs on day 10. Biochemical variables were similar among groups and times. CONCLUSIONS AND RELEVANCE: The administration of dipyrone in cats, when used in combination with tramadol, did not ensure better analgesia than tramadol alone. Dipyrone did not significantly affect biochemical variables and oxidative markers, despite minimal, clinically irrelevant, hematological differences between groups.

Interrelationships of chloride, bicarbonate, sodium, and hydrogen transport in the human ileum.

Using a triple-lumen constant perfusion system, the following observations were made in normal subjects. First, chloride, bicarbonate, and sodium were found to exhibit net movement across ileal mucosa against electrochemical gradients. Second, during perfusion with a balanced electrolyte solution simulating plasma, the ileum generally absorbed, but sometimes secreted fluid. A reciprocal net movement of chloride and bicarbonate was noted when sodium movement was zero. Increasing rates of sodium absorption were associated with decreasing bicarbonate secretion rates and finally bicarbonate absorption. Even when bicarbonate was absorbed ileal contents were alkalinized (by contraction of luminal volume). Third, net chloride movement was found to be sensitive to bicarbonate concentration in ileal fluid. For instance, chloride was absorbed from solutions containing 14 or 44 mEq/liter of bicarbonate, but was secreted when ileal fluid contained 87 mEq/liter of bicarbonate. Fourth, when chloridefree (sulfate) solutions were infused, the ileum absorbed sodium bicarbonate and the ileal contents were acidified. Fifth, when plasma-like solutions were infused, the potential difference (PD) between skin and ileal lumen was near zero and did not change when chloride was replaced by sulfate in the perfusion solution. These results suggest that ileal electrolyte transport occurs via a simultaneous double exchange, Cl/HCO2 and Na/H. In this model neither the anion nor the cation exchange causes net ion movement; net movement results from the chemical reaction between hydrogen and bicarbonate. No other unitary model explains all of the following observations: (a) human ileal transport in vivo is essentially nonelectrogenic even though Na, Cl, and HCO3 are transported against electrochemical gradients, (b) the ileum can secrete as well as absorb, (c) ileal contents are alkalinized during absorption of or during secretion into a plasma-like solution, and (d) the ileum acidifies its contents when sulfate replaces chloride. Data obtained with a carbonic anhydrase inhibitor support the proposed model.

PIP: Studies using a triple-lumen perfusion system in normal subjects were conducted to elucidate mechanisms of ileal electrolyte absorption. 4 primary observations were made in this study of interrelationships of chloride, bicarbonate, sodium, and hydrogen transport in human ileum: 1) chloride, sodium, and bicarbonate all exhibited net movement across ileal mucosa against electrochemical gradients; 2) when perfusion was performed in conjunction with administration of a balanced electrolyte solution stimulating plasma, the ileum generally absorbed, but sometimes secreted fluid; 3) net chloride movement was sensitive to bicarbonate concentration in ileal fluids; and 4) infusion of chloride-free (sulfate) solutions showed that the ileum absorbed sodium bicarbonate and that the ileal contents were acidified. A model to explain these findings suggests that ileal electrolyte transport occurs via a simultaneous double exchange, Cl/HCO3 and Na/H. In this model, net movement results from the chemical reaction between hydrogen and bicarbonate, not because of anion or cationic exchange.

Mechanism of bicarbonate absorption and its relationship to sodium transport in the human jejunum.

Using a constant perfusion technique, sodium and bicarbonate absorption was studied in human subjects. The following observations were made on sodium absorption from saline solution: (a) the rate of sodium absorption is markedly influenced by bulk water flow, (b) when net water flow is zero, sodium absorption is zero if there are no concentration gradients between plasma and lumen that favor net NaCl diffusion; and (c) the PD between abraded skin and jejunal lumen is near zero when saline is perfused and does not change with partial substitution of sulfate or bicarbonate for chloride. Based on these observations, we conclude that sodium absorption from saline is entirely passive in the human jejunum. On the other hand, in the presence of bicarbonate sodium is absorbed actively against electrochemical gradients. The mechanism of the link between bicarbonate and sodium absorption was studied in normal subjects and in 11 patients with pernicious anemia; the latter were chosen because they do not secrete gastric acid which can react with bicarbonate in the jejunal lumen. We observed that bicarbonate absorption (a) occurs against steep electrochemical gradients, (b) does not generate a potential difference between abraded skin and jejunal lumen, (c) is inhibited by acetazolamide, and (d) generates a high CO2 tension in jejunal fluid. These observations suggest that bicarbonate absorption is mediated by active hydrogen secretion, rather than by bicarbonate ion transport per se, and that the link between sodium and bicarbonate transport is best explained by a sodium-hydrogen exchange process.

PIP: In this study of bicarbonate and sodium absorption in the intestine, absorption in a 30-cm segment of intestine was studied by the Ingelfinger triple-lumen perfusion system, which involves perfusion of test solutions into the intestine and sampling of gut contents 10 and 40 cm beyond the infusion marker. Human subjects were used. Observations made from these experiments on the mechanism of bicarbonate absorption and its relationship to sodium transport in the jejunum from saline solutions include: 1) the rate of sodium absorption is influenced greatly by bulk water flow; 2) when net water flow is zero, sodium absorption is zero in the absence of concentration gradients betwee plasma and lumen; and 3) the potential difference between abraded skin and jejunal lumen is near zero when saline is perfused and does not change when sulfate or bicarbonate is partially substituted for the chloride. It is concluded that sodium absorption from saline is entirely passive in the human jejunum; in the presence of bicarbonate, sodium is actively absorbed against electrochemical gradients. This study also looked at the mechanism of the link between bicarbonate and sodium absorption. Normal subjects and 11 patients with pernicious anemia were studied. Bicarbonate absorption was found to 1) occur against steep electrochemical gradients; 2) not generate a potential difference between abraded skin and jejunal lumen; 3) be inhibited by acetazolamide; and 4) generate a high carbon dioxide tension in jejunal. These observations led to the conclusion that bicarbonate absorption is mediated by active hydrogen secretion rather than by bicarbonate ion transport per se, making the best explanation for the link between sodium and bicarbonate transport a sodium-hydrogen exchange process.

Stimulation of active and passive sodium absorption by sugars in the human jejunum.

The effects of glucose and fructose on water and sodium absorption in the human jejunum were compared to assess the relative contribution of active and passive sugar stimulation of sodium transport. The effect of fructose is assumed to be entirely passive, and the difference between the effects of fructose and glucose is assumed to be a measure of sugar-stimulated, active sodium absorption. Water and sodium movement with mannitol was the base line. Three sets of test solutions with differing sugar concentrations were studied. Fructose stimulated 66-100 per cent as much net sodium and water absorption as glucose. Fructose stimulated potassium absorption, whereas glucose stimulated potassium secretion. Urea absorption was stimulated by both sugars. Glucose and fructose stimulated sodium absorption when chloride was the major anion, but they had relatively little effect on net sodium movement when chloride was replaced by bicarbonate or sulfate. It is concluded that glucose stimulates passive and active sodium transport in the human jejunum. Stimulated active sodium absorption generates an electrical potential across the mucosa that causes sodium (and potassium) secretion and partly or completely nullifies the effect of active sodium transport on net sodium movement. Net sodium absorption sitmulated by glucose is mainly (66-100 per cent) the passive consequence of solvent flow. The accompanying anion determines the degree to which sugars stimulate sodium absorption (C1 greater than SO-4 greater than HCO3). The effects of bicarbonate and sugars on jejunal sodium absorption are not additive.

PIP: The carrier interaction and solvent drag components of sugar-stimulated sodium absorption were evaluated by comparing the effects of mannitol, fructose, and glucose on jejunal absorption of water, sodium, potassium, and urea. Using water and sodium movement with mannitol as the baseline, 3 sets of test solutions with differing sugar concentrations were studied. Fructose stimulated 66-100% as much net sodium and water absorption as glucose; in addition, fructose stimulated potassium absorption, whereas glucose stimulated potassium secretion. Both sugars stimulated urea absorption. When chloride was the major anion in the solution, glucose and fructose stimulated sodium absorption, however, the sugars had little effect on net sodium movement when chloride was replaced by bicarbonate or sulfate. From these observations it was concluded that glucose stimulates both passive and active sodium transport in human jejunum. Active sodium absorption generates an electrical potential causing secretion. Net sodium absorption stimulated by glucose was mainly the passive consequence of solvent flow. The effects of bicarbonate and sugars on jejunal sodium absorption were not additive.

Influence of augmented Hageman factor (Factor XII) titers on the cryoactivation of plasma prorenin in women using oral contraceptive agents.

Prolonged cold storage of plasma may induce the conversion of plasma prorenin (inactive renin) to renin. This phenomenon is exaggerated in oral contraceptive (OC) users; the titer of Hageman factor (HF, Factor XII) in OC users is higher than in nonusers. The present study relates these observations. The increment in plasma renin activity (PRA) during cold storage, as measured by generation of angiotensin I, correlated strongly with the initial plasma titer of HF. Increasing the HF titer of nonusers to that observed in OC users by addition of purified HF increased cold-induced PRA at least twofold, while reducing the plasma HF titer of OC users correspondingly decreased cold-induced PRA. Thus, in OC users, the enhanced conversion of plasma prorenin to renin during cold storage reflects the elevated plasma titer of HF.

PIP: Prolonged cold storage of plasma may induce the conversion of plasma prorenin (inactive renin) to renin. This phenomenon is exaggerated in oral contraceptive (OC) users; the titer of Hageman factor (HF, Factor 12) in OC users is higher than in nonusers. The present study relates these observations. The increment in plasma renin activity (PRA) during cold storage, as measured by generation of angiotensin I, correlated strongly with the initial plasma titer of HF. Increasing the HF titer of nonusers to that observed in OC users by the addition of purified HF increased cold-induced PRA at least 2-fold, while reducing the plasma HF titer of OC users correspondingly decreased cold-induced PRA. Thus, in OC users, the enhanced conversion of plasma prorenin to renin during cold storage reflects the elevated plasma titer of HF.

Hematological parameters on the effect of the jellyfish venom Cassiopea andromeda in animal models.

For the first time, we previously recorded an enormous population of the Cassiopea andromeda jellyfish that had increased dramatically from Bushehr coasts of Iran. The sub-acute toxicity of the jellyfish venom in rat organs was correspondingly carried out. The data presented in this paper relate to the in vivo and in vitro hematological effects of this venomous species of jellyfish venom.

The detection of alpha 1-fetoprotein in patients with viral hepatitis.

PIP: Alpha fetoprotein (AFP) was detected in sera (351 samples) of 128 patients with viral hepatitis by radioimmunoassay. 77 positive tests for AFP were obtained. These positive results were demonstrated on 1 or more samples taken from 40 (31%) of the 128 patients studied; the highest value obtained was 4400 ng/ml. Hepatitis B antigen (HBAg) was positive in 26/40 (65%) of patients in whom AFP was detected during the disease process. However, 58/88 (66%) who were seronegative for AFP also demonstrated HBAg in their sera. Chi-square analysis revealed no significant difference in occurrence of detectable AFP between HBAg seropositive and seronegative patients. Individuals seropositive for AFP had no statistically different concentration of the protein than patients seropositive or seronegative for HBAg. 24 patients' sera were tested serially over a 2-week period. Both the peak glutamic-pyruvic transaminase (GPT) and peak total bilirubin levels were in a higher range in those 10/24 patients seropositive (P .001) for AFP than in the 14/24 who were seronegative. Appearance of AFP was related to the severity of liver tissue destruction, as reflected by serum GPT. However, peak AFP levels were attained 5-16 days after peak serum GPT appeared in the circulation.^ieng

Early appearance of embryonic -globulin in rat serum during carcinogenesis with 4-dimethylaminoazobenzene.

PIP: Heptatic tumors were induced in male Donryo rats by feeding them a 4-dimethylaminoazobenzene diet to study the occurrence of serum alpha globulin (AG) in rats during carcinogenesis. AG was found in rat serum as early as the 3rd week after onset of feeding of the carcinogenic diet; this was designated the early-stage appearance. The embryonic protein was observed in 31 (76%) of 41 rats at the 6th week after diet introduction. Subsequently, concentrations of AG decreased, and by the 11th-12th week it disappeared from serum. After 13 weeks, the developmental protein reappeared in 27/33 rats, designated the last-stage appearance, and 26 of these animals developed hepatomas. Of the 22 rats in which AG appeared in the early stage, 20 (91%) developed hepatomas after 19 weeks. By the 6th week of the carcinogenic diet, the average serum level of AG was 2-4 mg/dl, but after 13 weeks, it reached 60-100 mg/dl, corresponding to the serum level of newborns. Since most of the rats in which AG appeared at the early stage developed hepatomas, the early appearance of the protein may be related to cancerization of liver cells; on the other hand, the early appearance of AG may simply reflect an acute liver lesion caused by the toxicity of 4-dimethylaminoazobenzene. All rats that developed hepatomas were AG positive.^ieng

Radioimmunoassay of rat 1 -fetoprotein.

PIP: A competitive binding radioimmunoassay for rat alpha 1 fetoprotein (AFP) was developed, using Sprague-Dawley rat amniotic fluid. The assay was approximately 20,000 times more sensitive than double-diffusion in agar for AFP detection; the assay threshold was 5 ng. Further purification of apparently pure (by immunodiffusion and immunization) radiolabeled AFP was required for the specific assay. An assay for a previously undetected contaminant(s) was used to check the purity of rat AFP isolated by isoelectric focusing to obtain the purified unlabeled AFP needed to establish the standard inhibition curve. All procedures are outlined.^ieng

Early appearance of serum -fetoprotein as a function of dosage of various hepatocarcinogens.

PIP: A previously reported early appearance of alpha fetoprotein (AFP) in rats fed 3'-methyl-4-dimethylaminobenzene (3-MDAB) which was induced before definite cancers were formed and disappeared on cessation of 3-MDAB administration was further investigated using different doses of 3-MDAB as well as other hepatocellular carcinogens and hepatotoxic agents. AFP was induced after 3 weeks of ingestion of diets with 600 ppm 3-MDAB and appeared after only 2 weeks when higher doses (900 and 1200 ppm) were used. Lower levels of 300 ppm 3-MDAB gave only a transient appaerance of AFP, beginning at the 5th week and remaining detectable for 3 more weeks, but 150 ppm did not induce at all. Immunosuppression with rat lymphocyte globulin extended for 1 week the time during which positive AFP titers were maintained upon cessation of 3-MDAB (600 ppm) intake. A transient appearance of AFP was found when rats were given the carcinogens dimethyl-4-dimethylaminoazobenzene (4-DMAA; 600 ppm), aflatoxin (AFB1; 4 ppm), N-2-fluorenylacetamide (N-2-FAA; 200 and 300 ppm), and N-hydroxy-N-2-fluorenylacetamide (N-OHFAA; 213 and 320 ppm). Lower doses of AFB1 (.2 and 2 ppm), N-2-FAA (150 ppm), N-OHFAA, and diethylnitrosamine (40 ppm) did not induce detectable AFP levels in serum nor did 2'-methyl-4-DMAA (600 ppm) and CCl4 (50 mg intraperitoneally twice a week). Apparently, high levels of liver carcinogens are required to induce the early appearance, within 2-5 weeks, of detectable AFP in serum.^ieng

Inhibition of antibody-complement-mediated killing of tumor cells by hormones.

Line 1, a chemically induced guinea pig hepatoma, is susceptible to killing by anti-Forssman immunoglobulin M antibody and guinea pig complement. When these tumor cells are pretreated at 37 degrees with 10(-4) to 10(-11) M concentrations of the polypeptide hormone insulin, with the catecholamine L-epinephrine-HCl, or with the glucocorticoid steroids hydrocortisone sodium succinate or prednisolone sodium succinate, the cells show a marked reduction in their suseptibility to killing by antibody and guinea pig complement; pretreatment at 0 degrees is ineffective. Similar results were obtained with another antigenically distinct guinea pig hepatoma (line 10) when tested with anti-Forssman immuno-globulin M or specific antitumor antibodies and human complement. The ability of the hormones to render the cells resistant is dependent on time, temperature, and hormone concentration. The effect of hormone treatment is maximal between 30 and 60 min and is reversible within 4 hr even in the continued presence of hormone. Treatment of line 1 cells with up to 10,000-fold greater concentrations of the less biologically active or inactive analogs, DL-epinephrine, beta-estradiol, testosterone, or proinsulin has no effect on the susceptibility of the cells to killing by antibody and guinea pig complement. The effect of hormone treatment is not due to a direct inactivation of bound or fluid-phase complement components by the hormones or to a decrease in the ability of the cells to bind complement-fixing antibody.

PIP: Various aspects of hormone treatment of tumor cells are reported; it is shown that following treatment with certain hormones, the cells are less susceptible to killing by antibody and complement. The diethylnitrosamine-induced guinea pig hepatoma, designated Line 1, is susceptible to killing by anti-Forssman immunoglobulin M (IgM) antibody and guinea pig complement (GPC) but not by specific antitumor antibody and GPC. The antigenetically distinct Line 10 hepatoma, when sensitized with either antibody, is susceptible to killing by human complement (HUC) but not by GPC. Strain 2 of Servall-Wright male guinea pigs were used. 2 antigenetically distinct diethylnitrosamine-induced hepatic tumors (ascites form), Lines 1 and 10, passed in Strain 2 guinea pigs, were collected and suspended in RPMI 1640-20% FCS. Toxicity assays were performed in VBS-gel. The hormones used were hydrocortisone sodium succinate, prednisolone sodium succinate, NSC9151, bovine insulin, L-epinephrine methyl ether HC1, DL-epinephrine, beta-estradiol, testosterone, pork insulin, chicken insulin, pork proinsulin, pork DAA insulin, and the A and B chains of pork insulin. Tumor cells were cultured in 10-ml volumes of RPMI 1640-20% FCS in plastic Petri dishes. After incubation, cell cultures were washed 5 times in VBS-gel and tested for their susceptibility to killing by antibody and complement. Rabbit antiserum to sheep Forssman antigen was prepared and stored at -20 degrees until used. Tumor specific rabbit Antilines 1 and 10 antisera were prepared and similarly stored. Results of tests show that Line 1 tumor cells incubated in a medium containing the polypeptide hormone, insulin, the catecholamine, L-epinephrine HCl, or the glucocorticoid steroids, hydrocortisone sodium succinate, or prednisolone sodium succinate were rendered resistant to killing byanti-Forssman IgM antibody and GPC. This effect was dependent on hormone concentration, temperature, and time. Effects were reversible. Similar results were obtianed with Line 10 cells under attack by specific antitumor and HUC or anti-Forssman antibodies. Less physiologically active analogs of the hormones did not have this effect. Tumor cells showed maximum resistance within 30-60 minutes of exposure to the hormones and reverted to the sensitive state within 4 hours. Resistance of the cells to killing was observed at 37 degrees but not at 0 degrees. It is concluded that the effect of hormone treatment was not due to a direct inactivation of bound or fluid-phase complement components by the hormones or to a decrease in the ability of the cells to bind complement-fixing antibody.

Alpha-fetoprotein in ontogenesis and its association with malignant tumors.

PIP: The major steps in development of ontogenesis and the role of alpha fetoprotein (AFP) synthesis are outlined. AFP is defined and its physiocochemical characteristics are described including methods of detection and identification. The liver and yolk sac of fetuses are shown as the principle sites of AFP synthesis in ontogenesis, and the dynamics of AFP in ontogenesis from the early embryonic period through midpregnancy to pregnancy termination to AFP shut-down in early postnatal period are displayed. AFP synthesis during regeneration of the liver provides the model for studying the nature of AFP production. The role of AFP in hepatocellular cancer receives a great deal of attention, focusing on the site of AFP synthesis in cancer of the liver; demonstration of AFP in blood of cases of hepatic cancer (and other diseases) by agar-gel precipitation; quantitative aspects of AFP production by liver tumors; and etiologic and pathogenic influences on AFP production by hepatomas. Clinical aspects of the diagnosis of liver cancer are reviewed. The occurrence of AFP with teratocarcinomas is remarked upon. The article's central objective was to emphasize the importance of basic research on AFP, especially the development of an accessible high-sensitivity test for use in broad epidemiological surveys. Experimental approaches to some immediate problems were formulated: 1) Is there any external factor controlling AFP synthesis and determining its intensity? 2) Is synthesis performed only by certain cell types or is AFP production inherent in any hepatocyte? 3) Is control of AFP synthesis accomplished by regulating the intensity of the process in individual cells or by involvement of a varying number of cells? And 4) is AFP synthesis in a tumor due to maintained ability of the stem tumor cell to differentiate or is it the result of dedifferentiation of the mature hepatocyte??^ieng

Estrogen therapy for severe persistent depressions in women.

Positive results are reported from a double-blind study of estrogen therapy administered to severely depressed, inpatient women who had failed to respond to various conventional treatments of depression. Large doses of oral conjugated estrogen were administered for a three-month period to 23 premenopausal and postmenopausal inpatient women. Placebos were administered for a comparable period to 17 similar patients. The posttreatment Hamilton ratings of depression were significantly reduced in the estrogen-treated group, but not in the placebo group. Possible physiological mechanisms are discussed. The risk-benefit ratio for estrogen therapy of depression in these patients was judged to be favorable. However, periodic endometrial biopsies are required to monitor the endometrial response of women receiving high doses of estrogens.

PIP: The effectiveness of estrogen therapy for alleviating severe depressions was investigated in a double blind study in which large doses of estrogen were administered to 15 premenopausal and 8 postmenopausal women and placebos were administered to 12 premenopausal and 5 postmenopausal women. The estrogens and placebos were administered over a three month period, and the women were blind rated each week by psychologists and psychiatrists using Hamilton rating scales for depression. There was a significant decline in the depression scores for the group treated with estrogens when compared to the placebo treated group. Considerable variation in the degree of improvement for the women in the estrogen treated group was observed. These variations were not related to age or to menstrual status but were significantly related to depression duration. Women with shorter histories of depression were more likely to show improvement than women with longer illness durations. Efforts were also made to understand the physiological mechanisms through which estrogen treatment contributed toward reducing depression. Previous studies revealed that decreased availability of norepinephrine at the central adrenegic receptor sites in the brain was related to the manifestation of depression while increased availability of norepinephrine at these sites was ralated to a manifestation of elation. Increased availability of norepinephrine has been shown to be related to an inhibition of monoamine oxidase activity (MAO), and estrogen, in turn, has been demonstrated to inhibit MAO activity. During the 3 month study period, 2 blood samples were obtained from the women every week and analyzed for MAO activity. All patients had elevated levels of plasma MAO activity prior to treatment. In estrogen treated patients, MAO levels declined significantly; MAO levels unexplicably increased for the placebo treated group. Reduced MAO activity was not, however, significantly correlated with lower depression ratings. In determining the risk/benefit ratio of estrogen therapy both the risk of developing endometrial cancer and the risks of life long disability and suicide stemming from severe depression should be considered. Tables show 1) mean depression ratings and average MAO activity levels before and after treatment for the estrogen and placebo treated groups and 2) degree of change in depression ratings before and after treatment for both groups of women.

Serological survey of HIV-1, HIV-2 and human T-cell leukemia virus type 1 for suspected AIDS cases in Ghana.

OBJECTIVE: To determine seroprevalence among suspected AIDS patients in Ghana in relation to clinical manifestations. MATERIALS AND METHODS: Blood samples and medical records were collected from 290 Ghanaian patients with suspected AIDS in 1990 and 1992. Seroprevalence of HIV-1, HIV-2 and human T-cell leukemia virus (HTLV-1) were investigated by the particle agglutination method, indirect immunofluorescence assay, the monoepitope enzyme-linked immunosorbent assay and Western blot. RESULTS: The specimens were classified into five serologic categories: 78 were HIV-1-positive (26.9%), 25 were HIV-2-positive (8.6%), 17 dual-positive (5.9%), 16 indeterminate (5.5%) and 154 seronegative (53.1%). No significant difference was found between the clinical symptoms of patients with HIV-1 and HIV-2 infection. Of the patients, 14 (4.8%) were HTLV-1-seropositive, of whom 11 were also HIV-positive, indicating a significant correlation between the two groups of viral infections (P < 0.01). However, there was no evidence of an increase in severity of symptoms in cases of dual infection with HTLV-1 and HIV. CONCLUSIONS: HIV-1 infection is now dominant in Ghana in contrast to our previous survey in 1986 which showed the dominance of HIV-2. The change in seroprevalence suggests that an HIV-1 epidemic has been developing in recent years in this country, where HIV-2 was originally endemic. A relatively high prevalence of dual-reactive specimens implies the existence of highly cross-reactive strains of HIV or frequent coinfection with HIV-1 and HIV-2 in the region. The large number of seronegative patients with clinically diagnosed AIDS raises the question of the inadequacy of AIDS definitions based on clinical manifestations only.

Patterns and predictors of high-risk sexual behavior in female partners of HIV-infected men with hemophilia.

OBJECTIVE: To characterize and quantify high-risk heterosexual activity in HIV-discordant couples. DESIGN: Analysis of cross-sectional and longitudinal questionnaire data from 217 HIV-negative female sexual partners of HIV-infected hemophilic men. METHODS: Comparison of prevalence rates of anal sex, oral sex, vaginal intercourse with or without condoms, and use of other contraceptives between 1985 and 1991. Logistic regression analysis of demographic, sexual and clinical variables to predict unprotected vaginal sex. Actuarial estimates of semi-annual relapse rates to unsafe sex. RESULTS: The proportion of women at low risk increased from 7 to 69% between 1985 and 1991, mainly because more women were using condoms during all acts of vaginal intercourse. Other contraceptive practices did not change during this time. The proportion engaging in oral or anal sex decreased (from 26 to 13% and 13 to 4%, respectively). Unprotected vaginal sex was more common among women who enrolled earlier, had less education, engaged in oral or anal sex, and among those whose partners had not had AIDS. Unprotected vaginal sex before enrollment was the strongest predictor of this high-risk activity during follow-up. Two-year rates of relapse to high-risk behavior were significantly higher among women who enrolled at high risk compared with those who enrolled at low risk (39 versus 8%, P = 0.005). CONCLUSIONS: Although high-risk sexual behavior became much less prevalent in this population between 1985 and 1991, many continued to have unprotected vaginal sex occasionally. Counseling efforts should target couples who have been the most sexually active or have less education, and should emphasize not only initial risk reduction but also maintenance of low-risk behavior.

PIP: This study sought to characterize and quantify the high-risk heterosexual activity in HIV-discordant couples. An analysis of cross-sectional and longitudinal questionnaire data from 217 HIV-negative female sexual partners of HIV-infected hemophiliac men were included in this study. There was a comparison of prevalence rates of anal sex, oral sex, vaginal intercourse with or without condoms, and use of other contraceptives between 1985-91. Logistic regression analysis of demographic, sexual, and clinical variables was used to predict unprotected vaginal sex. Actuarial estimates of semiannual relapse rates to unsafe sex were used. The proportion of women at low risk increases from 7 to 69% between 1985-91, mainly because more women were using condoms during all acts of vaginal intercourse. Other contraceptive practices did not change during this time. The proportion who engaged in oral or anal sex decreased from 26 to 13% and from 13% to 4%, respectively. Unprotected vaginal sex was more common among women who enrolled earlier, had less education, engaged in oral or anal sex, and among those whose partners had not had AIDS. Unprotected vaginal sex before enrollment was the strongest predictor of this high-risk activity during followup. 2-year rates of relapse to high-risk behavior were significantly higher among women who enrolled at high risk compared with those who enrolled at low risk (39 vs 8%, p=0.005). Although high risk sexual behavior become much less prevalent in this population between 1985-91, many continued to have unprotected vaginal sex occasionally. Counseling efforts should target couples who have been the most sexually active or have less education, and should emphasize not only initial risk reduction but also maintenance of low-risk behavior.