T cell induction of precursor B cells: Temporal separation of helper T cell functional activities

Summary The temporal relationships involved in T cell induction of immunoglobulin-secreting B cells have been studied by employing a pulse label technique, in vitro. It was shown that addition of rabbit thymocytes or splenic T cells to B cell-enriched splenocyte populations at the time of initiation of cultures resulted in a marked enhancement in induction of immunoglobulin-secreting cells. However, even a two-hour delay in the addition of thymus cells was sufficient to reduce substantially the extent of induction when measured 70 hours later. Besides this early requirement for thymocytes, a late requirement was also detectable. Thus, thymus cells and splenocyte populations upon being mixed, subsequent to being cultured separately for 72 hours, yielded a several-fold enhancement in [³H]-immunoglobulin secreted during the course of a 90-minute labeling period with [³H]-leucine. Moreover, both the early and late thymocyte effects were lost after treatment with anti-thymocyte serum and complement.The thymocyte-mediated enhancement of immunoglobulin secretion by splenocytes that occurs late in the induction process was detected with spleen cells cultured for two or three days but not with freshly-isolated splenocytes. Although the rate of appearance of extracellular immunoglobulins was markedly enhanced by fresh thymus cells, the rate of appearance of intracellular immunoglobulins in such spleen cells was unchanged. The secretion-stimulating (secretagogue) activity of thymocytes appeared to be specific in that thymus cells were without effect on the rate of secretion of serum albumin by liver cells.In regard to the induction of immunoglobulin-secreting cells, both B and T cell-enriched population's were sensitive to mitomycin C treatment performed before initiation of cell culture, indicating that not only B cells but also T cells undergo some form of differentiation or maturation prior to functioning in the induction of immunoglobulin-producing cells. It should be noted in this context that the late T cell requirement was unaffected by prior mitomycin C treatment of thymocytes. On the other hand, thymocytes heated at 60°C for 5 minutes did not enhance immunoglobulin secretion when added at any time and the late thymocyte requirement could not be replaced with medium in which thymocytes had been previously cultured.     

Involvement of glycoconjugates in insulin-receptor interactions Studies in liver plasma membranes of control and diabetic mice

The involvement of glycoconjugates in the insulin-receptor interactions in mouse liver is tested by digestions of membranes with various enzymes. Trypsin decreased the binding of [¹²⁵I]insulin to liver membranes. After digestion with β-galactosidase no “high affinity” receptor sites could be detected. The effects observed with plant lectins confirm the involvement of galactoconjugates in the insulin binding process. Sophora japonica and Ricinus communis lectins (with galactose specificity) and concanavalin A largely inhibit the binding process of insulin and those effects concern the “high affinity” receptor sites. Other lectins (wheat germ agglutinin, Dolichos) and enzymes (α-l-fucosidase, $\text{β-N-}\text{acetyl-hexosaminidase and neuraminidase}$) are without effect on insulin binding.Comparative studies performed on diabetic mouse liver membrane (KK mice), previously characterized by decreased number of insulin receptors, are in good agreement with qualitatively similar receptor sites in both non-diabetic (control) and diabetic mice. Effects of enzymes and lectins yielded same results as compared to control membranes. Plasma membrane proteins and glycoproteins in both types of mouse are indistinguishable with respect to enzymic and chemical analysis. Sodium dodecyl sulphate acrylamide gel electrophoresis shows identical patterns. Moreover, the decrease in the number of insulin receptors is easily reversed with diet restriction. These data are consistent with the similarity of receptor sites in control and diabetic liver membrane.     

Decreased binding of insulin to liver plasma membrane receptors in hereditary diabetic mice.

The interaction of insulin with its receptors was studied in liver plasma membranes of the young non-obese hereditary diabetic mouse (KK strain). Under identical conditions of preparation and incubation, the membranes of the KK mouse bind only 55-70% as much insulin per mg of protein as those of the control mouse (Swiss albino). Scatchard analysis suggests that this decrease in binding is due to a decrease in the number of receptor sites in the membrane of the diabetic mouse. However, the membranes of diabetic and control mice do not exhibit significant differences in hexosamine and sialic acid contents, enzyme activities, and protein and glycoprotein analysis. The decrease in insulin receptors in the KK mouse seems to correlate with the insulin resistance which they exhibit.     

Measurement of bile acid synthesis by 14CO2: the metabolism of propionyl CoA.

The relationship between 14CO2 evolution from the catabolism of [26 or 2714C] cholesterol to bile acids was studied in rats with biliary fistulae. When equal quantities of [26 or 2714C] cholesterol and [414C] cholesterol were administered, there was a significant linear relationship between 14CO2 expiration in the breath and [414C] bile acid excreted in the bile. Bile acid synthesis calculated as the ratio of 14CO2: molar specific activity of biliary cholesterol correlated highly with biliary bile acid excretion in the bile acid depleted rat. Phenobarbital, a known inducer of gamma-amino levulenic acid formation from succinyl CoA did not alter the relationship between the 14CO2 estimation of bile acid synthesis and biliary bile acid excretion, indicating that the relationship between [26 or 2714C] cholesterol side chain cleavage and 14CO2 formation was not altered. Phenobarbital, however, did cause a reduction in bile acid synthesis measured by 14CO2 evolution and by biliary bile acid excretion. The 14CO2 method underestimated bile acid excretion. 8.7% in untreated and phenobarbital treated rats respectively. Since 11% of the radioactivity which was expired as 14CO2 was isolated as bile acids, radioactivity cleaved as [1 or 314C] propionyl CoA may enter cholesterol-bile acid biosynthesis resulting in the underestimation of bile acid synthesis. To test whether radioactivity from propionyl CoA enters steroid biosynthesis [114C] propionate and [214C] propionate were given to untreated biliary fistula rats and the biliary lipids excreted in 60 hours were analyzed. Incorporation of radioactivity into cholesterol and bile acids was greater after the administration of [214C] propionate than after [114C] propionate than after [114C] propionate, suggesting that radioactivity from propionyl CoA may enter steroid biosynthesis by metabolic events in which the methylene and carboxyl carbon atoms are differentiated. Although the use of 14CO2 expiration from [26 or 2714C] cholesterol catabolism underestimates the rate of bile acid synthesis, it should have many applications because of the constant relationship between 14CO2 formation and cholesterol side chain cleavage.     

Observation and quantitation of metal-binding sites in the sex steroid-binding protein of human and rabbit sera using the luminescent probe terbium

The first direct evidence for specific metal-binding sites in pure human and pure rabbit sex steroid-binding protein (SBP) is obtained using the luminescent lanthanide terbium. Terbium, a probe for calcium sites in proteins, provided protection of the SBP steroid-binding activity in diluted human serum samples equivalent to that provided by calcium. Pure SBP, first treated with ethylenediaminetetraacetate, was dialyzed against buffer containing TbCl₃. After gel filtration to remove nonspecifically bound terbium, the protein was denatured in urea. The amount of protein-bound terbium was determined by luminescence enhancement of the lanthanide using the chelator dipicolinate, yielding four metal-binding sites per mole of dimer protein from both species.     

A novel early effect of concanavalin A on thymocytes.

Concanavalin A, added to freshly isolated rabbit thymocytes, markedly enhanced the extracellular appearance of non-immunoglobulin proteins. Time course studies revealed that the onset of enhancement occurred virtually without delay. The effect appeared to be restricted only to certain of the thymus-derived cells because thymocytes obtained from rabbits treated with hydrocortisone, as well as splenocytes derived from untreated rabbits essentially did not exhibit the enhancement. Stimulation by concanavalin A was specific in that pokeweed mitogen and lipopolysaccharide were without effect and also in that α-methyl-mannoside, but not galactose, abrogated the concanavalin A-mediated enhancement. Experiments with mouse thymocytes demonstrated that the cells which responded to concanavalin A were primarily cells that bear the θ-antigen on their surface (T-cells).     

Efficacy of different types of aerobic exercise in fibromyalgia syndrome: a systematic review and meta-analysis of randomised controlled trials

Introduction  

The efficacy and the optimal type and volume of aerobic exercise (AE) in fibromyalgia syndrome (FMS) are not established. We therefore assessed the efficacy of different types and volumes of AE in FMS.