Estradiol-17 beta and progesterone increase ovine uterine suppressor cell activity

We evaluated the regulation of ovine uterine (UT) suppressor cell activity by progesterone (P4), estradiol-17 beta (E2), and P4 + E2 in ovariectomized (OVX) ewes. Following 14 d of steroid injections, endometrial cells (designated as UT cells) were recovered postmortem, and unfractionated and fractionated cells were assessed for suppression of autologous phytohemagglutinin (PHA)-treated peripheral blood lymphocytes (PBL). Supernatants from cultured UT cells were also assessed for suppressor activity. In other experiments, UT cells recovered from nontreated OVX ewes were cocultured with PHA-treated PBL and varying concentrations (1 x 10(-11) to 1 x 10(-5) M) of each steroid preparation. Supernatants from separate cultures that contained UT cells and steroids were evaluated for suppressor activity. Uterine cells from control and steroid-treated ewes suppressed proliferative responses of PHA-treated PBL; however, suppressor activity of UT cells was greater (P < .05) for E2-treated than for control and P4-treated ewes. Uterine suppressor cells from steroid-treated ewes sedimented in Percoll within a density range of 1.002 to 1.056 g/mL. Uterine cells from all ewes released suppressor factor(s) into the culture medium; however, the activity of the supernatant from the cultured cells was not increased for the steroid-treated ewes. For cocultures that contained steroids and cultures that contained supernatant, suppressor activity of the UT cells was increased by specific concentrations of each steroid preparation. These findings demonstrate that reproductive steroids augment ovine UT suppressor cell activity.     

In vivo dexamethasone effects on neutrophil effector functions in a rat model of acute lung injury

1. In healthy male volunteers, the absorption, metabolite profiles and excretion of 14C-benidipine hydrochloride, a new Ca antagonist, were investigated after oral administration at a dose of 8 mg. 2. 14C-benidipine hydrochloride was rapidly absorbed, and the plasma concentration of radioactivity and unchanged drug reached a maximum of 71.2 ng eq./ml at 1.1 h and 2.56 ng/ml at 0.6 h respectively, and then declined bi-exponentially. The half-life in the elimination phase was 14.7 and 5.3 h respectively, AUC of unchanged drug was low, about 1% of that of radioactivity. 3. Five days after administration, 36.4% of the administered radioactivity was excreted in urine and 58.9% in faeces. 4. The metabolite profiles in plasma, urine and faeces were analysed by hplc. At 1 h after administration the predominant metabolites in plasma were M9 and M2, which accounted for 13.8 and 8.2% of the radioactivity respectively, whereas unchanged drug represented 1.2%. Predominant metabolites in urine 12 h after administration were M3 and M8, which accounted for 2.22 and 2.21% of the administered radioactivity respectively. Metabolites excreted in faeces 120 h after administration were very complex and poorly separated by hplc and could not be characterized: unchanged drug was not detected in the faeces.     

Intensive 50-week evaluation of glipizide administration in 50 cats with previously untreated diabetes mellitus

OBJECTIVE: To evaluate use of the oral hypoglycemic drug glipizide in diabetic cats. DESIGN: Prospective study. ANIMALS: 50 cats with recently diagnosed but untreated diabetes mellitus. PROCEDURE: Each cat received glipizide (5 mg, q 12 h) for 16 weeks. Medication was not given during the subsequent 16 weeks; then glipizide treatment was repeated. Each cat was evaluated prior to treatment and at 2, 4, 8, 12, and 16 weeks during each of the 3 phases: blood samples for serum glucose and insulin determinations were obtained every 2 hours, from 8 AM to 6 PM. A preprandial blood glycosylated hemoglobin percentage was determined for the first sample obtained at each visit. RESULTS: During the first 22 weeks of the study, diabetes worsened in 28 of the 50 cats, which then were disqualified from the study and treated with insulin. Of the remaining 22 cats that improved clinically, 7 had corresponding metabolic improvement in each diabetes-related parameter assessed and did not become hypoglycemic. Six of the 22 cats became hypoglycemic. Glipizide was discontinued, and diabetes did not recur. Serum glucose concentration did not improve in 6. Three cats had metabolic and clinical improvement during initial glipizide treatment, but had recurrence of the disease during repeated treatment; glipizide was discontinued and insulin was administered. None of the 50 treated cats died, and observed morbidity was mild and transient. Transient anorexia and vomiting were observed in 8 cats, and 4 became transiently icteric with abnormal liver enzyme activities. CLINICAL IMPLICATIONS: Trial use of glipizide is feasible in diabetic cats of owners who are unable or unwilling to administer insulin.     

The persistence of leptospiral agglutinins titers in human sera diagnosed by the microscopic agglutination test

Human serum albumin (HSA) is used in large amounts as an excipient in many biopharmaceutical formulation to prevent loss of the active ingredient through adsorption and/or degradation. Traditionally, iso-electric focusing has been used to demonstrate charge heterogeneity in HSA preparations. In an effort to develop new methods for the analysis of formulation components, a capillary zone electrophoresis method was developed for the analysis of HSA. Under initial separation conditions using untreated silica capillaries and 20 mM sodium phosphate, pH 6.0 as electrophoretic buffer, HSA migrated as a single peak. Addition of 1,4-diaminobutane allowed separation of several components which could be further resolved by varying the buffer pH. Optimal separation conditions were attained at 5 mM 1,4-diaminobutane and pH 8.5. The reproducibility of the separation conditions was verified by using capillaries from a different manufacturer. A comparative analysis of HSA preparations from different manufacturers provided evidence that the method may be used to qualitatively differentiate individual preparations. The analysis of rhEPO formulations, composed largely of HSA, showed levels of heterogeneity comparable to that of HSA preparations. Electrospray ionisation mass spectrometry (ESA-MS) was used as an independent method to confirm the heterogeneous nature of HSA.     

Rapid turnover of connexin43 in the adult rat heart

Remodeling of the distribution of gap junctions is an important feature of anatomic substrates of arrhythmias in patients with healed myocardial infarcts. Mechanisms underlying this process are poorly understood but probably involve changes in gap junction protein (connexin) synthesis, assembly into channels, and degradation. The half-life of the principal cardiac gap junction protein, connexin43 (Cx43), is only 1.5 to 2 hours in primary cultures of neonatal myocytes, but it is unknown whether rapid turnover of Cx43 occurs in the adult heart or is unique to disaggregated neonatal myocytes that are actively reestablishing connections in vitro. To characterize connexin turnover dynamics in the adult heart and to elucidate its potential role in remodeling of gap junctions, we measured Cx43 turnover kinetics and characterized the proteolytic pathways involved in Cx43 degradation in isolated perfused adult rat hearts. Hearts were labeled for 40 minutes with Krebs-Henseleit buffer containing [35S]methionine, and then chase perfusions were performed with nonradioactive buffer for 0, 60, 120, and 240 minutes. Quantitative immunoprecipitation assays of Cx43 radioactivity in 4 hearts at each time point yielded a monoexponential decay curve indicating a Cx43 half-life of 1.3 hours. Proteolytic pathways responsible for Cx43 degradation were elucidated by perfusing isolated rat hearts for 4 hours with specific inhibitors of either lysosomal or proteasomal proteolysis. Immunoblot analysis demonstrated significant increases ( approximately 30%) in Cx43 content in hearts perfused with either lysosomal or proteasomal pathway inhibitors. Most of the Cx43 in hearts perfused with lysosomal inhibitors consisted of phosphorylated isoforms, whereas nonphosphorylated Cx43 accumulated selectively in hearts perfused with a specific proteasomal inhibitor. These results indicate that Cx43 turns over rapidly in the adult heart and is degraded by multiple proteolytic pathways. Regulation of Cx43 degradation could play an important role in gap junction remodeling in response to cardiac injury.     

Effect of a serotonergic extrinsic modulatory neuron (MCC) on radula mechanoafferent function in Aplysia

The serotonergic metacerebral cells (MCCs) and homologous neurons in related mollusks have been extensively investigated within the context of feeding. Although previous work has indicated that the MCCs exert widespread actions, MCC modulation of sensory neurons has not been identified. We characterized interactions between the MCCs and a cell that is part of a recently described group of buccal radula mechanoafferents. The cell, B21, has a peripheral process in the tissue underlying the chitinous radula [the subradula tissue (SRT)]. Previous studies have shown that B21 can fire phasically during ingestive motor programs and provide excitatory drive to the circuitry active during radula closing/retraction. We now show that activity of B21 can be modulated by serotonin (5-HT) and the MCCs. Centrally, although a slow depolarization is typically recorded in B21 as a result of MCC stimulation, this depolarization does not cause B21 to spike. It can, however, increase B21 excitability enabling a pulse that was previously subthreshold to elicit an action potential in B21. B21 is in fact rhythmically depolarized during the radula closing/retraction phase of ingestive motor programs. Thus central effects of the MCCs on radula mechanoafferent activity are only likely to be apparent while B21 is receiving input from the feeding central pattern generator. Peripherally, radula mechanoafferent neurons can be activated 1) when a mechanical stimulus is applied to the biting surface of the SRT and 2) when the SRT contracts. MCC stimulation and 5-HT modulate B21 responses to both types of stimuli. For example, MCC stimulation and low concentrations of 5-HT cause subthreshold mechanical stimuli applied to the SRT to become suprathreshold. 5-HT and MCC stimulation also enhance SRT contractility. Peripheral effects of MCC activity are also likely to be phase dependent. For example, MCC stimulation does not cause B21 to respond to peripheral stimuli with an afterdischarge. Consequently, radula mechanoafferents are likely to be activated when food is present between the radula halves during radula closing/retraction but are not likely to continue to fire as opening/protraction is initiated. In a similar vein, MCC effects on the contractility of the SRT will only be apparent when contractions are elicited by motor neuron activity. SRT motor neurons are rhythmically activated during ingestive motor programs. Thus we have shown that radula mechanoafferent activity can be modulated by the MCCs and that this modulation is likely to occur in a phase-dependent manner.     

Point mutations in the beta chain CDR3 can alter the T cell receptor recognition pattern on an MHC class I/peptide complex over a broad interface area

To study how the T cell receptor interacts with its cognate ligand, the MHC/peptide complex, we used site directed mutagenesis to generate single point mutants that alter amino acids in the CDR3beta loop of a H-2Kb restricted TCR (N30.7) specific for an immunodominant peptide N52-N59 (VSV8) derived from the vesicular stomatitis virus nucleocapsid. The effect of each mutation on antigen recognition was analyzed using wild type H-2Kb and VSV8 peptide, as well as H-2Kb and VSV8 variants carrying single replacements at residues known to be exposed to the TCR. These analyses revealed that point mutations at some positions in the CDR3beta loop abrogated recognition entirely, while mutations at other CDR3beta positions caused an altered pattern of antigen recognition over a broad area on the MHC/peptide surface. This area included the N-terminus of the peptide, as well as residues of the MHC alpha1 and alpha2 helices flanking this region. Assuming that the N30 TCR docks on the MHC/peptide with an orientation similar to that recently observed in two different TCR-MHC/peptide crystal structures, our findings would suggest that single amino acid alterations within CDR3beta can affect the interaction of the TCR with an MHC surface region distal from the predicted CDR3beta-Kb/VSV8 interface. Such unique recognition capabilities are generated with minimal alterations in the CDR3 loops of the TCR. These observations suggest the hypothesis that extensive changes in the recognition pattern due to small perturbations in the CDR3 structure appears to be a structural strategy for generating a highly diversified TCR repertoire with specificity for a wide variety of antigens.     

Complementary surgical/interventional techniques for nonresective management of "inoperable" aneurysms: a second look

A review of the studies of the caries prevalence and periodontal health of patients with cleft lip and palate (CLP) revealed that only five investigations of caries prevalence in CLP patients included children. One reported that CLP children did not have a higher caries prevalence in the permanent dentition, whereas more recent studies have reported a higher caries prevalence in both the primary and permanent dentitions of CLP children than in those of noncleft children. However, there is wide variation in the teeth examined and the method of presenting data on caries prevalence. Six papers have reported on the periodontal health of adult CLP patients, and only one has done so on that of children. The adult CLP patients had poorer oral hygiene and more gingivitis, but there is no conclusive evidence that they have a higher risk of developing periodontal disease. No data on the oral hygiene of CLP children were available, but it has been emphasized that they have significantly more gingivitis than noncleft children, especially in the maxillary anterior teeth.