Comparative effects of a recombinant and a mutein type of granulocyte colony stimulating factor on the growth of Meth-A fibrosarcoma with 5-fluorouracil chemotherapy

The present study was designed to evaluate the effects of a recombinant human G-CSF (rhG-CSF) and a mutein G-CSF(KW-2228) on leucopenia and tumor growth in mice treated with 5-fluorouracil (5-FU). In normal mice, the number of leucocytes (white blood cell, WBC) reached the peak 12 hours after a single injection of either type of G-CSF and decreased to the normal level after 24 hours. Daily administration induced a continuous increase in the WBC count, however, administrations at intervals did not. Meth-A fibrosarcoma was subcutaneously inoculated into the backs of syngeneic BALB/c mice. The mice were treated with 5-FU alone or with G-CSFs. Chemotherapy with 5-FU alone resulted in leucopenia and an insignificant inhibition of tumor growth. The conjunctive administration of G-CSFs with 5-FU resulted in a significantly augmented inhibition of tumor growth, and leukopenia was not seen. This augmenting effect was more prominent with KW-2228.These results suggest that in 5-FU chemotherapy G-CSFs may be beneficial in restoring the number of leucocytes from leucopenic state and in augmenting the tumor inhibitory effect. Furthermore, KW-2228 may be more beneficial than the natural type rhG-CSF.     

Effect of substitution of troponin C in cardiac myofibrils with skeletal troponin C or calmodulin on the Ca2+- and Sr2+-sensitive ATPase activity

Troponin C was removed almost completely from the porcine cardiac myofibrils by the same extraction procedure using CDTA as that previously reported for the rabbit skeletal myofibrils (Morimoto, S. & Ohtsuki, I. (1987) J. Biochem. 101, 291-301), and the effects of substitution of troponin C in cardiac myofibrils with rabbit skeletal troponin C or bovine brain calmodulin were examined. While the ATPase activity of intact cardiac myofibrils or cardiac troponin C-reconstituted cardiac myofibrils was activated at only a little higher concentration of Sr2+ than Ca2+, the skeletal troponin C-substituted cardiac myofibrils, as well as intact rabbit skeletal myofibrils, required more than 10 times higher concentration of Sr2+ than Ca2+ for activation of the myofibrillar ATPase activity. However, the concentrations of Ca2+ and Sr2+ required for the activation of the ATPase activity of the skeletal troponin C-substituted cardiac myofibrils were both about 5 times higher than those of intact skeletal myofibrils. The skeletal troponin C-substituted cardiac myofibrils, as well as intact skeletal myofibrils, also showed higher cooperativity in the Ca2+-activation of the ATPase activity than intact or cardiac troponin C-reconstituted cardiac myofibrils. The ATPase activity of calmodulin-substituted cardiac myofibrils was activated at a several times lower concentration of Ca2+ or Sr2+ than that of calmodulin-substituted skeletal myofibrils, while the ratios of the concentration of Sr2+ to Ca2+ required for activation were almost the same in both cases.(ABSTRACT TRUNCATED AT 250 WORDS)     

Crystallization and preliminary X-ray studies on human epidermal growth factor

Single crystals of human epidermal growth factor have been prepared from a polyethylene glycol solution and characterized by X-ray diffraction. The crystals grow in a space group of P2(1) with unit cell dimensions of a = 32.7, b = 32.5, c = 22.3 A, and beta = 96.9 degrees. They contain a single molecule per asymmetric unit and show better diffraction than 2.5 A.     

Increase of (Ca2++Mg2+)-ATPase activity of renal basolateral membrane by parathyroid hormone via cyclic AMP-dependent membrane phosphorylation

Studies were made on the mechanism of the effect of parathyroid hormone (PTH) on the activity of (Ca2++Mg2+)-ATPase, a membrane bound Ca2+-extrusion pump enzyme from the basolateral membranes (BLM) of canine kidney (Km for free Ca2+ = 1.3 X 10(-7) M, Vmax = 200 nmol Pi/mg/min). At 1 X 10(-7) M free Ca2+, both PTH (10(-7)-10(-6) M) and cAMP (10(-6)-10(-4) M) stimulated (Ca2++Mg2+)-ATPase activity dose-dependent and their stimulatory effects were inhibited completely by 5 microM H-8, an inhibitor of cAMP-dependent protein kinase. PTH (10(-7) M) also caused 40% increase in 32P incorporation into the BLM and 5 microM H-8 inhibited this increase too. PTH (10(-7) M) was found to stimulate phosphorylation of a protein of Mr 9000 by cAMP dependent protein kinase and 5 microM H-8 was found to block this stimulation also. From these results, it is proposed that PTH stimulates (Ca2++Mg2+)-ATPase activity by enhancing its affinity for free Ca2+ via cAMP-dependent phosphorylation of a BLM protein of Mr 9000.     

Investigation of the specificity of O6-alkylguanine-DNA-alkyltransferase

Comparison of the abilities of alkylated RNA and DNA to serve as substrates for the O6-alkylguanine-DNA-alkyltransferase have been carried out. It was found that the O6-methylguanine in tRNA was much less active as a substrate for the protein than O6-methylguanine in double stranded DNA. The difference in rates of repair was such that it is unlikely that the alkyltransferase would act on RNA in vivo and, therefore, the reaction with RNA should not contribute towards the exhaustion of its repair capacity.     

A patient with hypogonadotropic hypogonadism successfully treated by long-term pulsatile administration of luteinizing hormone-releasing hormone

A male patient with hypogonadotropic hypogonadism has been treated by pulsatile administration lf luteinizing hormone-releasing hormone (LHRH) (20-25 micrograms, every 2 hours, sc) for 4 years 6 months. His plasma testosterone (T) concentration began to increase after 4 weeks of treatment and reached the normal range in week 5. He showed complete secondary sexual development after 1 year of treatment. His sperm count was normalized after 1 year of treatment. He was married after 29 months of therapy, and has a healthy male child. Blood type tests showed his paternity of the child. During the long duration of pulsatile LHRH therapy, his gonadotropin secretion has been stimulated by LHRH and his T level has been maintained with no observable side effects. There are no other reports of patients treated by pulsatile LHRH injection for such a long duration, but finding in this patient indicated that long-term pulsatile LHRH therapy is a useful and safe method for treatment of hypothalamic hypogonadotropic hypogonadism.     

Oxygen equilibrium study and light absorption spectra of Ni(II)-Fe(II) hybrid hemoglobins

Ni(II)-Fe(II) hybrid hemoglobins, in which hemes in either the alpha or beta subunit are substituted with Ni(II) protoporphyrin IX, have been prepared and characterized. Since Ni(II) protoporphyrin IX binds neither oxygen nor carbon monoxide, the oxygen equilibrium properties of the Fe subunit in these hybrid hemoglobins were specifically determined. K1 values, namely the equilibrium constants for the first oxygen molecule to bind to hemoglobin, agreed well for these hybrid hemoglobins with the K1 value of native hemoglobin A in various conditions. Therefore, Ni(II) protoporphyrin IX in these hybrid hemoglobins behaves like a permanently deoxygenated heme. Both Ne-Fe hybrid hemoglobins bound oxygen non-co-operatively at low pH values. When the pH was raised, alpha 2 (Fe) beta 2 (Ni) showed co-operativity, but the complementary hybrid, alpha 2 (Ni) beta 2 (Fe), did not show co-operativity even at pH 8.5. The light absorption spectra of Ni(II)-Fe(II) hybrid hemoglobins indicated that the coordination states of Ni(II) protoporphyrin IX in the alpha subunits responded to the structure of the hybrid, whereas those in the beta subunits were hardly changed. In a deoxy-like structure (the structure that looks like that observed in deoxyhemoglobin), four-co-ordinated Ni(II) protoporphyrin IX was dominant in the alpha (Ni) subunits, while under the conditions that stabilized an oxy-like structure (the structure that looks like that observed in oxyhemoglobin), five-co-ordinated Ni(II) protoporphyrin IX increased. The small change observed in the absorption spectrum of the beta (Ni) subunits is not related to the change of the co-ordination number of Ni(II) protoporphyrin IX. Non-co-operative binding of oxygen to the beta subunits in alpha 2 (Ni) beta 2 (Fe) accompanied the change of absorption spectrum in the alpha (Ni) subunits. We propose a possible interpretation of this unique feature.     

Fever in rats during normal and dehydrated conditions

The effect of endogenous pyrogen (EP, from rabbit) and endotoxin (Salmonella typhosa) on rectal temperature (Tre) was investigated in normal and dehydrated rats of both sexes. Intraperitoneal injection of either EP or endotoxin did not affect body temperature. In addition, no changes in Tre were observed when endotoxin was injected intravenously in normally hydrated male rats, but significant falls in Tre occurred in normal female rats. However, intravenous injection of EP produced fever in both sexes, but females generally showed smaller responses. A second intravenous injection of endotoxin, given 3 days after the first injection, always produced fever in normally hydrated rats. The pattern of this febrile response was monophasic. In contrast to the response in normal rats, intravenous endotoxin produced significant fevers with a biphasic pattern in dehydrated rats of either sex, but the febrile responses of male rats were greater than those of female rats. On the other hand, there were no significant differences between febrile responses to intravenous EP exhibited by normal and dehydrated animals. These results show that rats of both sexes possess physiological mechanisms capable of producing a fever following intravenous injections of EP.