The synthesis and localization of alternatively spliced fibronectin EIIIB in resting and thrombin-treated megakaryocytes

There are several species of alternatively spliced fibronectin (FN). One of these, FN EIIIB, is primarily present in embryonic and in proliferating and migrating cells and is believed to be important for cell maturation. We have studied the synthesis, localization, and secretion of this FN isoform in isolated guinea pig megakaryocytes, nonmegakaryocytic bone marrow cells, and platelets. There was 7.5 times more general FN in megakaryocytes than in nonmegakaryocytic cells based on the analysis of equivalent amounts of protein. FN EIIIB was detected by Western blotting in megakaryocytes but not in nonmegakaryocytic cells present in bone marrow. Neither megakaryocytes nor platelets secreted FN EIIIB, while megakaryocytes secreted 25.3% +/- 4.6% general FN and platelets secreted about 61% general FN in response to thrombin. Analysis of immunostained cells by confocal microscopy revealed that FN EIIIB had been redistributed to the surface of megakaryocytes in response to thrombin. Synthesis was studied by metabolic labeling, and megakaryocytes were shown to synthesize FN and FN EIIIB. Thus, megakaryocytes and platelets are among a small number of adult cells and tissues that synthesize and contain FN EIIIB. The expression of FN EIIIB on the megakaryocyte surface may influence migration and maturation.     

Interleukin-4 is a neutrophil activator

We investigated the ability of the lymphokine, interleukin-4 (IL-4), to function as a neutrophil (PMN) activator. IL-4 enhanced PMN-mediated killing of opsonized bacteria (by up to 91.6% at 3 units of IL-4; p less than 0.05). IL-4 was a weak secondary granule secretagogue and did not by itself generate a respiratory burst. However, IL-4 did increase in a dose-dependent fashion the respiratory burst mediated by the peptide formyl-methionyl-leucyl-phenylalanine (10(-7) mol/L). Maximal potentiation of PMN activity occurred at 100 units of IL-4 (6.3 nmol superoxide produced without IL-4 to 9.8 nmol at 100 units; p less than 0.01). Enhancement of the respiratory burst was not a generalized phenomenon, since IL-4 did not potentiate the respiratory burst mediated by either phorbol myristate acetate, calcium ionophore A23187, or zymosan-treated serum. Similarly, IL-4 potentiated the formyl-methionyl-leucyl-phenylalanine-stimulated secretion of both lysozyme (40.2%) and beta-glucuronidase (108.2%). Finally, IL-4 was demonstrated to enhance the ability of PMN to phagocytose sheep erythrocytes opsonized with rabbit IgG (by up to 94.2% at 30 units of IL-4). This increased phagocytosis correlated with the recruitment of a population of PMNs that did not phagocytose targets in the absence of IL-4. In conclusion, IL-4 enhanced neutrophil-mediated bactericidal activity. This increase may have occurred secondary to the stimulation of phagocytosis by IL-4 or by potentiation of degranulation and the respiratory burst.     

Young red blood cells; observations about the cytometric and morphologic alterations in the beginning of their life

Summary A recently described separation technique consisting of a combination of counterflow centrifugation and subsequent density (Percoll) scparation was tested for its ability to enrich red cell populations with young cells in comparison to either separation technique alone. The relative age of every fraction was determined by HbAlc measurements, resulting in the lowest HbAlc for the combination method. Conventional reticulocyte counting and floweytometric counting with thiazole orange indicated that in the youngest fractions the combination method showed the highest reticulocyte counts. There was a good correlation between manual and flowcytometric counting results. Radio-iron studies showed a two-fold enrichment with young cells in the fraction with the lowest HbAIc acquired by the combination technique in comparison to the other two methods. Cytometric measurements showed that the fractions with the lowest HbAlc were the ones with the highest MCV and MCH and the lowest MCHC. Besides loss of their RNA-material, young cells already seem to loose water and haemoglobin like older cells, resulting in a decrease of MCV and MCH and in increase in MCHC. It is concluded that combining counterflow centrifugation with subsequent density fractionation results in superior enrichment with young cells in comparison to the results of each method alone.     

Effects of high-intensity exercise on leptin and testosterone concentrations in well-trained males

Objective: A number of investigations have examined the effect of exercise on leptin concentrations, because leptin is associated with obesity, satiety, and reproductive function. High-intensity exercise is known to increase testosterone, an inhibitor of leptin. The objective of the study was to determine whether the leptin responses to a progressive, intermittent exercise protocol were related to serum testosterone concentrations. Most previous studies have examined leptin responses to low or moderately high exercise intensities. A second objective was to determine whether leptin responses were different than previous experiments using intermittent moderate and high-intensity exercise. Methods: Well-trained runners completed strenuous intermittent exercise consisting of treadmill running at 60, 75, 90, and 100% VO _2max and a subsequent resting control trial was also conducted. Results: There were significant increases in mean serum levels of leptin and testosterone with both quickly returning to baseline during recovery, but no relationship between the two hormones was found. After examining individual data for both hormones, it was discovered that subjects could be classified as leptin responders or nonresponders, whereas testosterone increased in all subjects. Responders had elevated serum leptin levels at baseline and exhibited increases after high-intensity exercise, whereas nonresponders did not show changes in leptin during exercise. Conclusions: Data suggest testosterone levels do not acutely affect leptin responses to exercise or 1-h of recovery. Moreover, varied leptin responses to intense exercise in comparable well-trained runners was observed and was associated with baseline leptin concentrations.     

MRI compatible MS2 nanoparticles designed to cross the blood–brain-barrier: providing a path towards tinnitus treatment

Fundamental challenges of targeting specific brain regions for treatment using pharmacotherapeutic nanoparticle (NP) carriers include circumventing the blood–brain-barrier (BBB) and tracking delivery. Angiopep-2 (AP2) has been shown to facilitate the transport of large macromolecules and synthetic nanoparticles across the BBB. Thus, conjugation of AP2 to an MS2 bacteriophage based NP should also permit transport across the BBB. We have fabricated and tested a novel MS2 capsid-based NP conjugated to the ligand AP2. The reaction efficiency was determined to be over 70%, with up to two angiopep-2 conjugated per MS2 capsid protein. When linked with a porphyrin ring, manganese (Mn²⁺) remained stable within MS2 and was MRI detectable. Nanoparticles were introduced intracerebroventricularly or systemically. Systemic delivery yielded dose dependent, non-toxic accumulation of NPs in the midbrain. Design of a multifunctional MRI compatible NP platform provides a significant step forward for the diagnosis and treatment of intractable brain conditions, such as tinnitus.     

Effect of estrogen on serum DHEA in younger and older women and the relationship of DHEA to adiposity and gender

This case-controlled study consisted of 2 parts. The objective of part 1 was to determine the relationship between DHEA, body mass index (BMI), and age in young males, young females, and postmenopausal (PM) females. Part 2 examined the effects of estrogen on DHEA by analyzing the relationship between DHEA and age in young females on and off oral contraceptives (OCs) and PM females on and off estrogen or hormone replacement therapy (ERT/HRT). The study was performed at the Obstetrics and Gynecology Clinic, Texas Tech Health Sciences Center-Amarillo, Exercise Physiology Laboratory at Southeastern Louisiana University, and Woman's Health Research Institute, Woman's Hospital, Baton Rouge, LA. Part 1 groups consisted of: (1) young males between the ages of 18 to 40 years; (2) normally cycling females off OCs, ages 18 to 40 years; and (3) PM females older than 40 years not receiving ERT/HRT. Part 2 groups consisted of: (1) normally cycling females on OCs, ages 18 to 40 years;, (2) normally cycling females off OCs, ages 18 to 40 years; (3) PM females 50 years or older not receiving ERT/HRT; and (4) PM females 50 years or older receiving ERT/HRT. The main outcome measure was serum DHEA concentrations. For part 1, there were significant (P <.05) inverse relationships between DHEA and age for young males; young females, off OCs; PM females, no ERT/HRT r = -.44, -.26, and -.25, respectively. There were no significant relationships between DHEA and BMI for any of the groups. DHEA concentrations were significantly higher in young males than young females even after accounting for age. For part 2, DHEA concentrations were significantly higher in young females off OCs compared with young females on OCs, and significantly higher in PM women off ERT/HRT than those on ERT\HRT. There were significant inverse relationships between DHEA and age for young females and PM females on and off ERT/HRT. From these findings, we conclude that there is an inverse relationship between DHEA and age for young males, young females off OCs, and PM females, no ERT/HRT. No relationship between BMI and DHEA was observed in these same 3 groups. These results agree with previous findings in young men, but differ from previous findings in obese young females. The data also suggest that estrogen treatment (OCs and ERT/HRT) suppresses DHEA concentrations in premenopausal and PM females, and that DHEA declines with age in PM females regardless of estrogen treatment.     

Impact of the selective serotonin reuptake inhibitor citalopram on insulin sensitivity, leptin and basal cortisol secretion in depressed and non-depressed euglycemic women of reproductive age.

INTRODUCTION: Major depression in women of reproductive age may be accompanied by multiple endocrine and metabolic disturbances, which, in turn, may affect reproductive functioning. Enhanced cortisol synthesis, impaired leptin production and diminished insulin sensitivity have been reported in some depressed populations. We sought to determine whether an 8-week administration of citalopram would have an effect on these endocrine factors in a group of euglycemic depressed and non-depressed women of reproductive age. MATERIALS AND METHODS: Fourteen depressed and 18 non-depressed women (diagnosed by structured clinical interview) aged between 18 and 45 years completed an 8-week study. All depressed women were treated with citalopram and non-depressed subjects randomized to citalopram or no treatment in an open label cohort study. An oral glucose tolerance test with insulin levels was performed at baseline and at the end of the 8-week trial. Weight, blood pressure, fasting serum cortisol, fasting serum leptin and Beck Depression Inventory were assessed at baseline, 2, 4 and 8 weeks. RESULTS: Citalopram significantly improved depressive symptoms and Beck Depression Inventory scores in the depressed cohort. Cortisol production was higher in depressed women but did not diminish with citalopram therapy over 8 weeks. Indices of insulin sensitivity and leptin production were similar between depressed and non-depressed women and did not change despite citalopram therapy. CONCLUSION:. Insulin sensitivity in moderately depressed women of reproductive age does not differ from that in a similar group of non-depressed women. Insulin sensitivity, cortisol secretion and leptin production do not change significantly in depressed women following an 8-week course of citalopram despite substantial improvement in depression scores.