Thyroid-stimulating immunoglobulin bioassay using cultured human thyroid cells.

Modifications are described in the cultured thyroid cell cAMP assay for TSH which make it suitable for the measurement of thyroid-stimulating immunoglobulins. Comparison was made between this assay and two others measuring cAMP responsiveness in human thyroid tissue, namely the thyroid slice and thyroid plasma membrane adenylate cyclase assays, all performed with the same tissue sample. Of immunoglobulin G (IgG) samples from 7 unselected patients with untreated hyperthyroidism associated with Graves' disease, 5 produced significant stimulation of cAMP content in cultured thyroid cells when compared to pooled normal IgG. None of these 7 produced a statistically significant increase in thyroid slice cAMP content when assayed in triplicate, the same replicate number used in the cultured thyroid cell assay. Similarly, none of the same Graves' IgG samples produced significant stimulation (vs. control IgG) in the membrane adenylate cyclase assay, in which sensitivity to TSH stimulation was very poor. With a scaled-down modification of the assay using microtiter wells and acetylation to enhance detection of cAMP in the RIA, significant TSI activity was observed in 15 of 18 (83%) IgG samples from patients with untreated Graves' disease. The data indicate the excellent sensitivity and precision of the thyroid cell cAMP assay, as well as its convenience.

Specific suppression of graft-versus-host responsiveness by incubation of donor lymphoid cells with a solubilized membrane fraction of host lymphoid cells.

BALB/c spleen cells were incubated with a solubilized membrane fraction (SMF) prepared from spleen and thymus cells of (BALB/c x C3H/He)F1 or (BALB/c x A/J)F1 hybrid mice. Cells incubated with (BALB/c x C3H/He)F1 SMF produced less graft-versus-host (GVH) splenomegaly in (BALB/c x C3H/He)F1 hosts than did untreated BALB/c cells. The reduction of GVH splenomegaly was specific, inasmuch as the GVH activity of (BALB/c x C3H/He)F1 SMF-treated and untreated cells was similar in (BALB/c x C57BL)F1 hosts, and BALB/c cells treated with (BALB/c x A/J)F1 SMF showed no alteration of GVH activity in either (BALB/c x C3H/He)F1 hosts or (BALB/c x C57BL) F1 hosts. The time course of splenomegaly did not differ for SMF-treated and untreated cells. Donor cells that were labeled with tritiated adenosine and treated with (BALB/c x C3H/He) F1 SMF produced a reduction in the amount of label appearing in (BALB/c x C3H/He)F1 host spleens but not in (BALB/c x C57BL)F1 host spleens. Mechanisms which could account for the ability of SMF to cause specific reductions in both GVH activity and host spleen labeling are discussed.

Effects of 14 days of spaceflight and nine days of recovery on cell body size and succinate dehydrogenase activity of rat dorsal root ganglion neurons.

The cross-sectional areas and succinate dehydrogenase activities of L5 dorsal root ganglion neurons in rats were determined after 14 days of spaceflight and after nine days of recovery. The mean and distribution of the cross-sectional areas were similar to age-matched, ground-based controls for both the spaceflight and for the spaceflight plus recovery groups. The mean succinate dehydrogenase activity was significantly lower in spaceflight compared to aged-matched control rats, whereas the mean succinate dehydrogenase activity was similar in age-matched control and spaceflight plus recovery rats. The mean succinate dehydrogenase activity of neurons with cross-sectional areas between 1000 and 2000 microns2 was lower (between 7 and 10%) in both the spaceflight and the spaceflight plus recovery groups compared to the appropriate control groups. The reduction in the oxidative capacity of a subpopulation of sensory neurons having relatively large cross-sectional areas immediately following spaceflight and the sustained depression for nine days after returning to 1 g suggest that the 0 g environment induced significant alterations in proprioceptive function.

Myonuclear number and myosin heavy chain expression in rat soleus single muscle fibers after spaceflight.

The effects of 14 days of spaceflight on myonuclear number, fiber size, and myosin heavy chain (MHC) expression in isolated rat soleus muscle fiber segments were studied. Single soleus muscle fibers from rats flown on the Spacelab Life Sciences-2 14-day mission were compared with those from age-matched ground-based control rats by using confocal microscopy and gel electrophoresis. Spaceflight resulted in a significant reduction in the number of fibers expressing type I MHC and an increase in the number of fibers expressing type IIx or IIa MHC. Space-flight also resulted in an increase in the percentage of fibers coexpressing more than one MHC and in the reexpression of the neonatal isoform of MHC in some fibers. Fiber cross-sectional area was significantly reduced in pure type I MHC-expressing fibers and in fibers coexpressing type I+II MHC but not in fibers expressing one or more type II MHC in the flight rats. The number of myonuclei per millimeter was significantly reduced in type I MHC-expressing fibers from the flight rats but was not significantly different in type I+II and type II MHC-coexpressing fibers. Fibers expressing neonatal MHC were similar in size to control fibers but had significantly fewer myonuclei per millimeter than flight fibers not expressing neonatal MHC. In type I MHC-expressing fibers, the reduction in fiber cross-sectional area was greater than the reduction in myonuclear number; thus the average cytoplasmic volume per myonucleus was significantly lower in flight than in control fibers. The reduction in both myonuclear number and fiber size of fibers expressing type I MHC after 14 days of spaceflight supports the hypothesis that changes in the number of myonuclei may be a contributing factor to the reduction in fiber size associated with chronic unloading of the musculature.

Spaceflight effects on beta-adrenoceptor and metabolic properties in rat plantaris.

Effects of 14 days of spaceflight on beta-adrenoceptor (beta-AR), mitochondrial enzyme activities, and fiber type composition were studied in plantaris muscles of male adult Sprague-Dawley rats. The beta-AR was analyzed in cross sections by quantitative autoradiography. The maximum binding capacity (Bmax) of beta-AR was significantly lowered (approximately 29%) after flight, but the recovery was not completed within 9 days in 1-G environment. Because the dissociation constant remained unchanged, it is suggested that the changes in the Bmax were caused by the alteration of receptor number. The activities of succinate dehydrogenase (SDH) measured in whole homogenates were subnormal (approximately -24%) in muscles sampled approximately 5 h after flight but they were normalized during 9 days of recovery. The percent composition of fiber types and beta-hydroxyacyl CoA dehydrogenase (HAD) activity did not change significantly due to spaceflight. It is suggested that the spaceflight-induced decrease of the Bmax of beta-AR in plantaris was accompanied by a lowered activity of a mitochondrial inner-membrane enzyme SDH but not a matrix enzyme HAD.

Modulation by thymus-derived (T) cells of thyroid cell-stimulated prostaglandin E release by human peripheral blood mononuclear cells.

Cultures of plastic-adherent, human peripheral blood mononuclear cells generated prostaglandin E (PGE). Culture of the adherent cells (predominantly monocytes) with human thyroid cells enhanced PGE accumulation in the medium, although to a lesser degree than occurs with unseparated blood mononuclear cells. Recombination of the adherent cells with monocyte-depleted, nonadherent cells restored both basal and thyroid cell-stimulated PGE generation to the levels seen with unseparated cells. Thymus-derived (T) cells, obtained by rosetting with sheep erythrocytes, similarly enhanced both the adherent cell basal PGE production as well as the increased PGE accumulation that occurs in the presence of thyroid cells (50-120% augmentation by the T cells). Significant augmentation of thyroid cell-stimulated PGE release by 10(5) adherent cells occurred with the addition of as few as 5 x 10(4) T cells. Culture medium transfer experiments and separation of cell types during culture by a semipermeable membrane provided evidence against the possibility that the adherent cells were releasing a factor that stimulated T-cell PGE generation or that T cells were releasing a factor that enhanced adherent cell PGE generation. The results suggest instead that this T-cell effect requires direct contact with the adherent cells. These data demonstrate the importance of human T cells in the release by adherent cells of PGE, a mediator of suppressor function by some immune cells.

Influence of spaceflight on succinate dehydrogenase activity and soma size of rat ventral horn neurons.

Succinate dehydrogenase (SDH) activities and soma cross-sectional areas (CSA) of neurons in the dorsolateral region of the ventral horn at the L5 segmental level of the spinal cord in the rat were determined after 14 days of spaceflight and after 9 days of recovery on earth. The results were compared to those in age-matched ground-based control rats. Spinal cords were quick-frozen, and the SDH activity and CSA of a sample of neurons with a visible nucleus were determined using a digitizer and a computer-assisted image analysis system. An inverse relationship between CSA and SDH activity of neurons was observed in all groups of rats. No change in mean CSA or mean SDH activity or in the size distribution of neurons was observed following spaceflight or recovery. However, there was a selective decrease in the SDH activity of neurons with soma CSA between 500 and 800 microns2 in the flight rats, and this effect persisted for at least 9 days following return to 1 g. It remains to be determined whether the selected population of motoneurons or the specific motor pools affected by spaceflight may be restricted to specific muscles.

Comparison of the response of motoneurons innervating perineal and hind limb muscles to spaceflight and recovery.

The succinate dehydrogenase (SDH) activities and cell body sizes of motoneurons in the dorsomedial (DM) region of the ventral horn at the lower portion of the L5 and the L6 segmental levels of the rat spinal cord were determined following 14 days of spaceflight and after 9 days of recovery on Earth and compared with those in the retrodorsolateral (RDL) region of the ventral horn at the same segmental levels. No changes in the mean SDH activity of motoneurons in the DM region were observed following spaceflight or after recovery. However, a decrease in the mean SDH activity of motoneurons with cell body sizes between 500 and 900 microm(2) in the RDL region was observed following spaceflight and after recovery. These data indicate that moderate-sized motoneurons in the RDL region, which are most likely associated with the hind limb musculature, were responsive to the microgravity environment. In contrast, the motoneurons in the DM region associated with the perineal muscles (associated with predominantly fast, low-oxidative muscles which are recruited for relatively brief periods at high activation levels and have no load-bearing function at 1G) were not affected by microgravity.