Effects of short-term high dose intake of evening primrose oil on plasma and cellular fatty acid compositions, α-tocopherol levels,and erythropoiesis in normal and Type 1 (insulin-dependent) diabetic men

Summary In addition to their usual diet, nine Type 1 (insulin-dependent) diabetic men and ten male control subjects took 20 g d,ga-tocopheryl acetate enriched evening primrose oil (14.45 g 182c,6, 1.73g 183c,6, 400 mg d,-tocopheryl acetate) daily for one week. At start, diabetic patients had more 140, 150 and 18 2c,6, and less 160, 161c,7, 181c,7, 183c,6, 203c,9, 203c,6, 204c,6 and 226c,3 in plasma, erythrocytes and/or platelets. Furthermore, they had lower 161c,7/160, 181c,7/160, and 204c,6/203c,6 ratios and a higher 203c,6/183c,6 ratio. In diabetic patients, -tocopherol levels in erythrocytes were lower, whereas those in plasma were normal. In both groups, oil intake changed fatty acid profiles. Most markedly, 203c,6 increased, whereas the ratios 203c,6/ 183c,6 and 204c,6/203c,6 decreased. 204c,6 increased in control subjects, but not in diabetic patients. Erythrocytes and platelets responded differently in their fatty acid profiles, -tocopherol rose in plasma and, although less for diabetic patients, in erythrocytes. In diabetic patients as well as in control subjects, erythrocyte count, haemoglobin level, mean corpuscular haemoglobin content and concentration increased and glycosylated haemoglobin percentage decreased without an apparent decline in blood glucose levels. Plasma -thromboglobulin and platelet factor 4 decreased, especially in diabetic patients. In conclusion, diabetic patients had abnormal fatty acid patterns, suggesting an impaired 9, 6 and 5 desaturation and an enhanced chainelongation, and had lower erythrocyte a-tocopherol levels; and short-term high dose intake of evening primrose oil increased 203c,6 in both groups, but 204c,6 only in control subjects, gave fatty acid responses which were different for erythrocytes and platelets, enhanced erythropoiesis, and lowered indices of in vivo platelet activation.     

Effect of polyinosinic-polycytidylic acid on chemically induced tumorigenesis by methylcholanthrene in mice

Summary Polyinosinic-polycytidylic acid administered intraperitoneally inhibits the formation of chemically induced tumors by methylcholanthrene in mice. The experiments show that poly (IC) is an effective suppressor of tumor formation when given simultaneously with the cancerogenic compound, or soon thereafter (before 4 weeks). Once the tumorigenesis was started (after 8 weeks), poly (IC) treatment becomes ineffective.The mechanism of inhibition of tumor formation by poly (IC) was studied by measuring the immune response of treated mice. Mice treated with methylcholanthrene alone exhibit a 50% inhibition of the immune response towards sheep red blood cells. Animals injected with poly (IC) after the methylcholanthrene treatment did not show any significant change. However, a pretreatment with poly (IC) causes a complete reversal of immunosuppression caused by methylcholanthrene.

---

Zusammenfassung Polyinosin-Polycytidylsäure (Poly IC), intraperitoneal verabreicht, hemmt die Bildung von chemisch induzierten Tumoren durch Methylcholanthren in Mäusen. Die Versuche zeigen, daß Poly (IC) ein wirksamer Hemmstoff der Tumorbildung ist, wenn es gleichzeitig mit Methylcholanthren oder bald danach (vor Ablauf von 4 Wochen) gegeben wird. Wenn die Tumorgenese einmal begonnen hat (nach 8 Wochen), wird die Poly (IC)-Behandlung unwirksam.Der Hemmungsmechanismus der Tumorbildung durch Poly (IC) wurde durch Messung der Immunantwort von behandelten Mäusen untersucht. Nur mit Methylcholanthren behandelte Mäuse zeigen eine 50%ige Hemmung der Immunantwort in Gegenwart von Schaferythrocyten. Behandelt man die Tiere zuerst mit Methylcholanthren und anschließend mit Poly (IC), so bleibt die Immunantwort unbeeinflußt. Ändert man diese Reihenfolge, indem das Poly (IC) vor Methylcholanthren eingespritzt wird, so wird die immunsuppressive Wirkung des Methylcholanthrens vollkommen aufgehoben.

     

Different ratio of myosin heavy chain isoforms in arterial smooth muscle of spontaneously hypertensive rats

Summary The relative proportion of the two putative heavy chains of smooth muscle myosin (MHC1 and MHC2) was determined in the caudal and femoral arteries of spontaneously hypertensive rats (SHR) and normotensive (WKY) rats at 16 weeks of age. The heavy chain polypeptides with M_r 204000 and 200000 were resolved electrophoretically under denaturing conditions in porous polyacrylamide gels. Both proteins reacted strongly with a monoclonal antibody (2C4) to smooth muscle MHC. In caudal arteries the ratio of MHC1/MHC2 was 3.11 in SHR rats compared with 1.81 in WKY rats (p<0.005) and similarly in femoral arteries, 2.81 vs 1.51 (p<0.001). In the portal vein there was no significant difference, 1.71 vs 1.51. The possibility that the higher MHC ratio in the SHR is the genetically mediated defect in arterial smooth muscle cells leading to the hypertension is discussed as an alternative to the elevated systemic blood pressure causing the altered MHC ratio.     

Effect of β-adrenergic stimulation on free fatty acid content in subepicardial and subendocardial layers of the dog heart in situ. Separation and assay by capillary gas chromatography

Summary The effects of -adrenergic stimulation produced by an infusion of isoproterenol (1 g·kg^–1 min^–1, 30 min) were studiedin situ in the anaesthetized dog placed under a total cardiopulmonary bypass. Samples of the subepicardial and the subendocardial layers were homogenized separately prior to the extraction and methylation of free fatty acids (FFA). Gas chromatography on Carbowax 20 M capillary columns was used for the quantitation of myristic (C 140), palmitic (C 160), palmitoleic (C 161), stearic (C 180), oleic (C 181), linoleic (C 182), and arachidonic (C 204) acids.Within 5 min, isoproterenol decreased the tissue content of FFA significantly. The decrease was more pronounced in the endocardial layer where the FFA concentration reached its minimum at the 5th or the 15th min. In the epicardial layer, all the FFA reached their minimal concentration at the 30th min of the isoproterenol infusion. In both layers, lactate content remained unchanged at 5 and 15 min and rose at the 30th min only and content in phosphorylated compounds (ATP, creatine-phosphate—CP) did not show any significant variation during the -stimulation period. A significant correlation was found between the chronotropic effect of isoproterenol and the reduction of FFA concentration.With the technical assistance of Agnès Bacconin.     

Large-bowel cancer in the young: A national survival study

Large-bowel cancer in young patients is reported to be a more aggressive and advanced disease at presentation and is believed to be associated with a relatively poor prognosis. Of 2420 patients registered in New Zealand (1968 to 1970), 131 were under 40 years of age and 2289 were over 40 years of age. The annual average incidence of treatable colorectal cancer in patients under 40 years of age was 2.36 per 100,000 and 82.93 in patients over 40 year of age. There were predominantly more females in both age groups with colonic tumors, 5044 (femalemale), and 759652 (femalemale). The rectal tumor male-to-female ratio of 10.68 in those over 40 years of age was reversed in those under 40 years of age (12.08). There was no significant difference in the subsite distribution of colorectal cancers between the two groups. There was a higher proportion of Stage 1 tumors in those under 40 years of age and a correspondingly higher proportion of Stage 2 tumors in those over 40 years of age. The overall crude and relative five-year survival rates for patients under 40 years of age were both 60 percent, whereas the crude rate for older patients was 42 percent, with a corresponding relative rate of 53 percent. Ten-year survival rates were generally higher in younger patients. From this study, there was no evidence to suggest that younger patients (less than 40 years old) with colorectal cancer had worse prognoses and did not survive as long as older patients (40 years and over).Poster presentation at the meeting of The American Society of Colon and Rectal Surgeons, Anaheim, California, June 12 to 17, 1988.     

Chlamydia trachomatis serology in ankylosing spondylitis

Summary Demonstration of chlamydial antibodies in patients with ankylosing spondylitis (AS) could show an etiological roel of Chlamydia trachomatis in this condition. We studied serum specimens from 50 HLA-B27 positive patients with AS (Group I), 34 HLA-B27 positive patients with other rheumatic diseases (Group II), 67 HLA-B27 positive healthy blood donors (Group III) and 37 healthy untyped blood donors. (Group IV). Measured by an immunoperoxidase assay (IPA) chlamydial IgA (titre 120) was more prevalent in the HLA-B27 positive persons than in the healthy controls not selected for HLA-group (Groups I+II+III vs IV: p<0.02). Chlamydia trachomatis IgA-IPA containing sera also had specific IgG-IPA antibodies (180) in 29 (96%) out of 30 sera from HLA-B27 positive individuals and controls. Conversely, 45% of specific IgG-positive (180) AS sera, 27.7% sera in Group II, 39.4% Group III sera vs. 11.1% of sera in Group IV had concomitant chlamydial IgA (120). The differences in the prevalence of specific IgA were statistically significant: Group I vs. IV: p<0.01; Group III vs. IV: p<0.05 and Gr. I+II+III vs. IV: p<0.05. Our data suggest an enhanced antibody production against Chlamydia trachomatis among the HLA-B27 positive individuals whether they have AS or are healthy.     

Gewinnung von Granulozyten zur Transfusion mit dem IBM-Blutzellseparator von gesunden Spendern

Zusammenfassung Durch Variation der mit dem IBM-Zellseparator zur Gewinnung von Leukozyten gebräuchlichen Methoden konnte gezeigt werden, daß die Antikoagulierung mit einer Kombination von Heparin und ACD, die Verwendung von Neoplasmagel^® sowie die Reduzierung des extrakorporalen Blutvolumens die besten Voraussetzungen zur Gewinnung von Granulozyten sind. So konnten von einem gesunden Spender in 6 h im Mittel (n=5) 1,6x10¹⁰ Granulozyten isoliert werden bei einem Reinheitsgrad von Granulozyten: Lymphozyten: Thrombozyten: Erythrozyten=11,211151. Die Ergebnisse konnten darüber hinaus durch Erhöhung der Granulozytenausgangszahl beim Spender mit Prednisolon gesteigert werden auf eine mittlere Ausbeute (n=5) von 5,6x10¹⁰ bei einem Reinheitsgrad von Granulozyten: Lymphozyten: Thrombozyten: Erythrozyten=10,23,240. Die Viabilität der isolierten Granulozyten wurde geprüft durch Untersuchung ihres Verhaltens nach Markierung mit DF³²P und autologer Transfusion. Gegenüber nicht isolierten, im Vollblut markierten Granulozyten zeigte sich beiobne Prednisolon isolierten Granulozyten eine Verminderung, beimit Prednisolon isolierten Granulozyten eine Erhöhung der Transfusionseffektivität.

---

Summary By varying the common methods for separating leukocytes with the IBM cell separator the anticoagulation with a combination of Heparin and ACD, the use of Neoplasmagel^® as well as the reduction of the extracorporal blood volume have shown to be the best conditions to separate granulocytes. From 5 healthy donors a mean yield of 1.6x10¹⁰ granulocytes could be obtained during six h with a degree of purity of granulocytes: lymphocytes: thrombocytes: erythrocytes=11.211151. These results could be improved by increasing the count of granulocytes in the donor's blood with prednisolone in another 5 donors to a mean yield of 5.6x10¹⁰ with a degree of purity of granulocytes: lymphocytes: thrombocytes: erythrocytes=10.23.240. The viability of separated granulocytes was proved by investigating their circulation after labeling with³²P and autologous transfusion. Comparing with granulocytes which were not separated but labeled in fresh whole blood the effectivity of transfusion was diminished when separated granulocytes were transfused without stimulation with prednisolone and was increased when separated granulocytes were transfused after stimulation with prednisolone.

---

---

Studie im Rahmen des Assoziationsvertrages EURATOM-GSF No. 031-64-I BIAD und des Sonderforschungsbereiches 37 der Universität München.     

A prospective study of local recurrence after resection and low stapled anastomosis in 183 patients with rectal cancer

Local recurrence is the most serious complication of anterior resection for rectal cancer, usually occurring during the first two years after surgery. Over a five-year period, from 1981 to 1986, 183 patients underwent anterior resection for rectal carcinoma at the Surgery Ward of the University of Ferrara. Patients were followed for two years postoperatively. All operations were performed with staplers and classified according to Dukes, with 43 cases of Dukes' A; 83 cases of Dukes' B; and 57 cases of Dukes' C. In the first 24 months after surgery, the tumor recurred locally in 44 of the 183 patients (24 percent. Dukes' stage, grading distal resection margin, and histopathologic differentiation of the distal rectal ring left in the stapler after anastomosis were assessed to determine a prognostic indicator for the recurrence of the tumor. The stage:recurrence ratio was as follows: A, 1 (2 percent); B, 21 (25 percent); and C, 22 (39 percent). The grading:recurrence ratio was: G1, 1351 (25 percent); G2, 24110 (22 percent); and G3, 722 (32 percent). The ratio between distal rectal resection margin and recurrence was: 0 to 2 cm, 1527 (56 percent); 2 to 4 cm, 1674 (22 percent); and over 4 cm, 1382 (15 percent). Histopathologic examination of the distal rectal ring was negative for all patients. These data confirm the direct relationship between class and local recurrence and indicate histologic grade and distal resection margin as significant prognostic parameters only when interpreted in the light of staging.     

Hypomagnesaemia in Type 2 (non-insulin-dependent) diabetes mellitus is not corrected by improvement of long-term metabolic control

Summary Low levels of magnesium have frequently been reported in diabetes mellitus especially in poorly controlled Type 1 (insulin-dependent) diabetic patients. Furthermore hypomagnesaemia might contribute to insulin resistance in Type 2 (non-insulin-dependent) diabetes. As the influence of improved metabolic control on plasma magnesium levels is unknown in Type 2 diabetic patients we studied magnesium plasma levels in 50 patients 1) before, 2) one and 3) three months after the initiation of insulin therapy or intensified treatment with oral hypoglycaemic agents. Magnesium plasma levels were measured by a colorimetric method and were significantly reduced in diabetic patients compared to healthy control subjects (0.79±0.01 mmol/l vs 0.88±0.01 mmol/l; p<0.0001). Metabolic control was significantly improved as documented by reduced HbA_1C levels in both insulin-treated patients or the patients on oral hypoglycaemic agents (p<0.003). However, plasma magnesium levels remained unchanged during the follow-up in the insulin-treated group (10.79±0.02 mmol/l; 20.81±0.02 mmol/l; 30.79±0.01 mmol/l) as well as in the patients on oral hypoglycaemic agents (10.79±0.03 mmol/l; 20.78±0.02 mmol/ l; 30.84±0.04 mmol/l). This study shows that even marked improvement of glycaemic control does not correct hypomagnesaemia in Type 2 diabetes. We conclude that hypomagnesaemia might be related to the insulin-resistant state and that possible beneficial effect of chronic magnesium administration should be evaluated in these patients.     

In vivo and in vitro test for growth potential of liver cells from rats during early stage of hepatocarcinogenesis by 3′-methyl-4-dimethylaminoazobenzene

Summary A rapid increase in the fraction of small liver cells was observed in the liver of rats during the early stage of hepatocarcinogenesis by 3-methyl-4-dimethylaminoazobenzene (3-Me-DAB). The change in cell population was represented by the decrease in glucose-6-phosphatase activity and by the increase in number of -glutamyltranspeptidase-positive cells. When DNA synthesis of liver cells from rats fed 3-Me-DAB was measured by autoradiography in primary culture, it began to increase 2 weeks after the start of the carcinogen feeding, reaching a plateau level after 3 weeks. Liver cells from rats fed 3-Me-DAB for 2 weeks or over demonstrated a remarkable resistance to the cytotoxic effect of the carcinogen (0.24 mM) in primary culture. Furthermore, liver cells from rats fed 3-Me-DAB for 3 weeks or over proliferated in the presence of the carcinogen in primary culture. When liver cells from 3-Me-DAB-fed and control rats were transplanted into syngeneic rat spleens, the former cells proliferated more vigorously than did the latter. The growth potential of liver cells from 3-Me-DAB-fed rats tended to be enhanced with time in the carcinogen feeding. Hepatocellular carcinomas developed in the host spleens implanted with liver cells from a rat fed 3-Me-DAB for 8 weeks. As described above, liver cells from rats fed 3-Me-DAB demonstrated much greater proliferative ability than normal control cells in vivo and in vitro.Abbreviations used HCC hepatocellular carcinoma - 3-Me-DAB 3-methyl-4-dimethylaminoazobenzene - GGT -glutamyltranspeptidase     

Colonic mucosal T lymphocytes in ulcerative colitis: Expression of CD7 antigen in relation to MHC class II (HLA-D) antigens

T-cell subsets and their activation state were examined by double-label immunofluorescence of cryostat tissue sections of the colon from 21 patients with ulcerative colitis (UC) and 30 histologically normal controls. Expression of MHC class I (HLA-A, B, C) and class II (HLA-D) antigens was studied in parallel. In the normal colonic mucosa, the CD4CD8 ratio in the epithelial compartment approximated 11, and in the lamina propria, 2.551. Of the CD8⁺ (cytotoxic/suppressor) subset, approximately half did not express the CD5 pan-T marker in either compartment. Virtually no Leu 8⁺ cells were observed, implying that the CD4⁺ subset consisted of helper, rather than suppressor-inducer cells. Classical markers of T-cell activation (CD25, HLA-D) and proliferation were absent, and strong expression of the CD7 immunostimulation marker was approximately equal in both CD4 and CD8 subsets. The epithelium was uniformly negative for class II antigens, but positive for class I. In UC, there were no significant alterations in CD4CD8 ratios in either compartment, and there were no changes with respect to phenotype of the subsets. In 11 of 19 patients (mainly with total colitis), enterocytes were HLA-D⁺. In this HLA-D⁺ group, there was an increase in the percentage of CD4⁺ cells coexpressing CD7; this difference was significant (P<0.02) in the lamina propria. Increased expression of CD7 was also found by the CD6⁺ T cell subset (P<0.05). These results suggest that class II expression is mediated by immunostimulated T helper cells in UC, with consequences for antigen presentation and maintenance of the chronic inflammatory state.HLA-D is used as a generic term for class II major histocompatibility complex (MHC) gene products (HLA-DR, DP, DQ) unless specified otherwise.     

Analysis of lymphocyte subsets in patients with aplastic anemia before and during immunosuppressive therapy

Summary To define the contribution of T-lymphocyte subsets in the development of aplastic anemia (AA), T-cell subpopulations including T cells, T cells, and TCS1-positive T cells, were analyzed by cytophotometry in the peripheral blood (PB) and bone marrow (BM) of patients with AA before and after 6 weeks of therapy with anti-lymphocyte globulin (ALG), methylprednisolone, and cyclosporin A (CSA). In nine patients with AA a significant decrease of PB- and BM-derived T cells was observed after 6 weeks of therapy as compared with normal controls. At diagnosis, the CD4/CD8 ratio in PB and BM of the patients did not differ from the ratio in the control population; however, a reversed ratio (< 1) was present in PB as well as in BM after weeks of therapy. Interestingly, lymphocytes expressing the T-cell receptor (TCR) were significantly decreased both before (PB 1.2±0.1%; BM 0.8±0.1%) and after 6 weeks of therapy (PB 0.7±0.1%; BM 0.7±0.1%) as compared with healthy controls (PB 2.4±0.2%; BM 2.3±0.2%). However, the proportion of the -T-cell subpopulation expressing the TCS1 phenotype was markedly increased before (PB 42±3.5%; BM 31±3%) and especially after 42 days of therapy (PB 77±12%; BM 45±2%) as compared with that in normal subjects (PB 19±2%; BM 9.7±0.8%). At present, follow-up is under evaluation to correlate these findings with hematological response. The pathophysiological significance of the observed alterations within the T-cell subsets and especially the T-cell populations will require further functional analyses, in particular since TCS1-positive T cells exhibit autoimmunological capacity.Presented at the annual meeting of the German Society for Hematology and Oncology, 4–7 October 1992, Berlin     

Comparative and combined effects of transforming growth factors α and β, interleukin-1 and interferon-γ on rheumatoid synovial cell proliferation,glycolysis and prostaglandin E production

Summary Enhanced cell proliferation, glycolysis and prostaglandin E production are all characteristic features of rheumatoid synovial tissue. The interrelationships of these three cellular parameters have been examined using rheumatoid synovial fibroblasts and their responses to specific cytokines in vitro. Transforming growth factor (TGF) caused a more than threefold increase in synovial cell proliferation whilst transforming growth factor (TGF), interleukin-1 (IL-1) and interferon- (IFN-) produced only marginal changes. The combined addition of IL-1 with TGF resulted in an enhanced proliferative response comparable with that produced by TGF. Glycolysis, estimated by glucose utilisation and measurements of the glycolytic regulatory metabolite fructose 2,6-bisphosphate was significantly stimulated by TGF, IL-1 and IFN-, but less so by TGF. Prostaglandin E production was significantly increased by IL-1 to an extent much greater than that produced by TGF or TGF, although the combined addition of IL-1 with either TGF or resulted in a synergistic increase in PGE production, a response partly diminished by the addition of IFN-. These findings suggest that the extent to which a cytokine stimulates glycolysis is not consistently related to its mitogenicity, and that cytokine combinations which stimulate high levels of PGE production (a growth inhibitor) will not necessarily be associated with a reduced rate of cellular proliferation in cultured, adherent, rheumatoid synovial fibroblasts.     

Triplication (/ααα<Superscript>Anti3.7</Superscript>) or deletion (-α<Superscript>3.7</Superscript>/) association in Argentinian β-thalassemic carriers

Prevalence of alpha gene triplication or deletion in -thalassemia carriers was studied in 109 unrelated individuals in Rosario, Argentina. In different populations -^3.7 allele presents a higher prevalence than ^anti3.7; thus, -thalassemia associated with -thalassemia is more frequently observed. Nevertheless, this event was detected in only one case (0.9%), while the association with alpha triplication was present in two subjects (1.8%).     

Long-Term Clinical Remission Induced by Corticosteroid Withdrawal Therapy (CSWT) in Patients with Chronic Hepatitis B Infection: A Prospective Randomized Controlled Trial—CSWT with and Without Follow-Up Interferon-α Therapy

The effectiveness of corticosteroid withdrawal therapy (CSWT), with or without follow-up interferon- (IFN-), has not been reported for HBe antigen (HBeAg) -positive patients with chronic hepatitis B. We conducted a prospective randomized controlled trial in 42 patients with HBeAg- and HBV-DNA-positive chronic hepatitis B (HBV genotype C: 38 patients) to assess the possible additive effect of follow-up IFN- after CSWT compared with CSWT alone. HBeAg seroconversion rates in the CSWT-alone and the combination group were 11.1% vs 11.8% at 24 weeks, 27.8% vs 12.5% at 52 weeks, 33.3% vs 18.8% at 76 weeks, and 38.9% vs 18.8% at 104 weeks, respectively. The final HBeAg seroconversion rates after CSWT alone were twice those following combination therapy. We conclude that CSWT alone is a very short-term treatment of just three weeks that may be more effective for long-term clinical remission than CSWT followed by IFN- in Japanese genotype C-dominant hepatitis B patients.     

The effect of dbcAMP on the lysolecithin induced hemolysis of calf and adult cattle erythrocytes

Summary The calf erythrocytes have an increased sensitivity against lysolecithin as compared to their adult counterparts. 10^–3M dbcAMP increases the hemolysis induced by 5g of lysolecithin in 0.15 M NaCl containing 10 mM phosphate buffer (pH 7.4). By increasing the level of phosphate buffer (75 mM) in the incubation mixture, 10^–3M dbcAMP decreases the hemolysis induced by 5g of lysolecithin. These data suggest a dual effect exerted by dbcAMP: the relatively labilizing or stabilizing effect prevails as a function of exogenous inorganic phosphate level.10-⁶M dbcAMP also has a relative protective effect against lysolecithin.The combined addition of cAMP (10^–3) and theophyllin (10^–4M) does not stabilize the membrane.By increasing the level of lysolecithin to 20g/ml the stabilizing effect of dbcAMP disappears.DbcAMP (10^–3) as well as cAMP (10^–3M) and theophyllin (10^–4M) have a minimum increasing effect on hemolysis in the absence of lysolecithin, too.

---

Abbreviations dbcAMP N⁶-20-dibutyryladenosine 35-monophosphate - cAMP cyclic 35-adenosine monophosphate     

Gaucher disease (type 1): Physical and kinetic properties of liposomal and soluble ‘acid’ β-glucosidase

Acid -glucosidase of human spleen, from either normal controls or patients with type 1 (adult) Gaucher disease, was incorporated into phosphatidylcholine liposomes. The non-incorporated (soluble) Gaucherenzyme had a higher apparent molecular weight than had the corresponding control. Liposomal acid -glucosidase prepared from Gaucher-spleen was more thermostable than was the corresponding normal enzyme; it was also stimulated by acidic lipids to a much lesser extent. The results suggest that the genetic mutation in type 1 (adult) Gaucher disease has multiple effects on the glycoprotein form of acid -glucosidase.     

Differences of cellular composition and adhesion molecule expression in “leukemic” as compared with “normal” human long-term bone marrow cultures

Summary Human long-term bone marrow cultures (HLTBMCs) were established with bone marrow samples collected from 15 patients with acute myeloid leukemia (AML) and compared with HLTBMCs from eight healthy volunteers. During 6 weeks of culture, the cellular composition of HLTBMCs was quantitatively studied. The cells of the HLTBMCs were divided into three main categories: fibroblasts, macrophages, and other cells (endothelial cells, hematopoietic cells and undefined cells). HLTBMCs derived from healthy volunteers demonstrated a very consistent development. The number of fibroblasts increased during culture and the number of macrophages decreased, resulting in a steady state after 3 weeks of culture. In contrast, HLTBMCs derived from patients with AML showed a strikingly different pattern of irregular development and a steady state was not reached under our conditions. The APAAP technique was used to demonstrate expression of adhesion molecules. VLA2, VLA5, VLA6, LFA1, Mac1, p150/95, ₂-chain, HCAM, ICAM1, NCAM, and VCAM1 were more expressed on normal as compared with leukemic bone marrow stromal cells, although this reached significance only for ₂-chain and NCAM. VLA1, 3, and 4 were expressed in a higher percentage on leukemic stroma (not significant). More expression was seen on normal as opposed to leukemic macrophages for the adhesion molecules tested, except for VLA5. The differences reached significance for the majority of molecules tested. It is concluded that striking differences exist in cellular composition and adhesion molecule expression between HLTBMCs from healthy individuals and those from patients with AML. This may have an impact on the pathogenesis of AML.     

Dietary supplementation of undernourished rats with soy or safflower oil: Effects on myelin polyunsaturated fatty acids

Undernourished suckling rats were administered, by gastric intubation, either soy oil (which is rich in both linoleic and linolenic acids) or safflower oil (which is rich in linoleic acid but deficient in linolenic acid) to determine (1) if dietary supplementation would offset the hypomyelination characteristic of the undernourished, developing brain and (2) to compare myelin fatty acids in normal, undernourished, and oil-supplemented rats. Myelin recovery was not increased by supplementation with either oil. The proportions of C₂₂₄ and C₂₂₆ fatty acids were reduced in myelin of the undernourished rats. Undernourished rats supplemented with either soy or safflower oil had higher than normal proportions of polyunsaturated fatty acids (C₂₀₄ and C₂₂₆). The triene-tetraene ratio in the oil-supplemented rats was lower than in normal controls, indicating that the oil-supplemented rats were not deficient in essential fatty acids. No significant differences were observed between the oil-supplemented groups.     

Preferential inhibition of DNA polymerasesα,δ, andε from Novikoff hepatoma cells by inhibitors of cell proliferation

DNA polymerases , and from normal regenerating rat liver and Novikoff hepatoma cells were purified about 300-fold, characterized, and checked for sensitivity towards drugs known to inhibit cell proliferation. Characterization included (a) identification of associated proteins, (b) measurement of physicochemical constants (including sedimentation coefficients, diffusion coefficients, calculation of relative molecular masses), (c) quantification of catalytic activities using specific DNA primer templates (K _m values) and specific inhibitors (K _i values), and (d) discrimination between DNA polymerases from normal cells and those from malignant cells using inhibitors of cell proliferation. (a) DNA primase associated with DNA polymerase , and 3–5 exonuclease accompanying DNA polymerases and had similar activities. (b) Comparison of physicochemical and catalytic properties of DNA polymerases from both sources revealed similarities but also some important differences. Sedimentation and diffusion coefficients of DNA polymerases and from malignant cells differed significantly. (c) The DNA-binding domain of DNA polymerases and from hepatoma cells was altered sinceK _m values, determined with several specific DNA primer-templates, were higher. Furthermore, dNTP-binding sites of DNA polymerases from malignant cells; when probed with specific inhibitors (aphidicolin, butylphenyl-dGTP, carbonyldiphosphonate, and dideoxy-TTP) showed significantly lowerK _i values, indicating lower affinity to deoxyribonucleoside 5-triphosphates. (d) Sixteen drugs representative of various modes of interaction with DNA and protein were chosen. Dose/response experiments were performed and the concentration at which the polymerizing activity was reduced to 50% was calculated (K ₅₀ values). Preferential inhibition of DNA polymerases ,, and from Novikoff hepatoma cells was found for: the intercalating drugs doxorubicin, daunorubicin, amsacrine, mitoxantrone, quinacrine and ethidium bromide, the minor-groove binders distamycin and netropsin, the ATPase-blocking agents novobiocin and coumamycin, and the topoisomerase I inhibitors camptothecin and topotecan. When the sensitivity of polymerases and was measured using poly(dA·dT) as a primer-template, the preferential inhibition of the enzymes from malignant cells was even more pronounced. Drugs known to trap the DNA-topoisomerase-II complex, etoposide, nalidixic acid, teniposide, and merbarone did not affect DNA polymerases irrespective of the source. Since the majority of the inhibitors used, particularly intercalators and minor-groove binders, act by modification of the primer-template, inhibition of DNA synthesis must have occurred through weakening of non-covalent bonds between DNA and catalytic polypeptides. Consequently, preferential inhibition of DNA polymerases from malignant cells seems to be indicative of abnormally diminished binding of the enzymes to their primer-templates. This effect may be caused by conformational alterations in polymerases from malignant cells which affect the DNA binding domains. Similarly, changes in physicochemical and kinetic constants are indicative of alterations of dNTP-binding domains.Abbreviations butylphenyl-dGTP N ²-(p-n-butylphenyl)-2-deoxyguanosine 5-triphosphate - dideoxy-TTP 2,3-dideoxyribosylthymine 5-triphosphate - HEPES 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid - etoposide 4-demethylepipodophyllotoxin-9-(4,6-O-ethylidene--D-glucopyranoside) - amsacrine 4-(9-acridinylamino)methanesulfon-m-ansidide - mitoxantrone 1,4-dihydroxy-5,8-bis{2-[(2-hydroxyethyl)amino]ethyl} amino)-9,10-anthracenedione - merbarone 5-(N-phenylcarboxamido)-2-thiobarbituric acid - PCNA proliferating-cell nuclear antigen - teniposide 4-demethylepipodophyllotoxin-9-(4,6-O-thionylidine--d-glucopyranoside) - topotecan (S)-10-[(dimethylamino) methyl]-4-ethyl-4,9-dihydroxy-1H-pyrano[3, 46, 7] indolizino[1, 2-b]quinoline-3,14(4H, 12H)-dione hydrochloride