Ionophore-mediated uptake of ciprofloxacin and vincristine into large unilamellar vesicles exhibiting transmembrane ion gradients

A new method, based on the ion-translocating properties of the ionophores nigericin and A23187, is described for loading large unilamellar vesicles (LUVs) with the drugs vincristine and ciprofloxacin. LUVs composed of distearoylphosphatidylcholine/cholesterol (DSPC/Chol) (55:45 mol/mol) or sphingomyelin (SPM)/Chol (55:45 mol/mol) exhibiting a transmembrane salt gradient (for example, internal solution 300 mM MnSO4 or K2SO4; external solution 300 mM sucrose) are incubated in the presence of drug and, for experiments involving divalent cations, the chelator EDTA. The addition of ionophore couples the outward movement of the entrapped cation to the inward movement of protons, thus acidifying the vesicle interior. External drugs that are weak bases can be taken up in response to this induced transmembrane pH gradient. It is shown that both nigericin and A23187 facilitate the rapid uptake of vincristine and ciprofloxacin, with entrapment levels approaching 100% and excellent retention in vitro. Following drug loading, the ionophores can be removed by gel exclusion chromatography, dialysis, or treatment with biobeads. In vitro leakage assays (addition of 50% mouse serum) and in vivo pharmacokinetic studies (in mice) reveal that the A23187/Mn2+ system exhibits superior drug retention over the nigericin/K+ system, and compares favorably with vesicles loaded by the standard DeltapH or amine methods. The unique features of this methodology and possible benefits are discussed.     

Establishment and characterization of three cell lines from Aedes triseriatus (Diptera: Culicidae)

The objectives of this study were to develop sex-, age-, and body size-specific nomograms and partition values for upper and lower limits of M-mode echocardiographic aortic root measurements derived from a large population-based cohort. The study sample consisted of 1433 male and 1816 female participants in the Framingham Heart Study and Framingham Offspring Study who were normotensive and free of clinically apparent heart disease at the baseline examination. Aortic root measurements were obtained by M-mode echocardiography by a leading-edge to leading-edge technique. The relations of age and measures of body size with aortic root dimensions were evaluated with sex-specific correlations and multiple stepwise linear regression analyses. Age was the most important determinant of aortic root size in both men and women in the multivariable regression models. Models with age and body surface area yielded R2 values of 0.214 in men and 0.222 in women. Models with age and height yielded lower R2 values of 0.136 in men and 0.181 in women. Thus aortic root dimensions vary widely with the age, sex, and body size of individuals. Sex-specific reference nomograms of aortic root dimensions in relation to age and body size (body surface area or height) are presented to facilitate the detection of abnormalities of aortic root size.     

Is urine leukocyte esterase test a useful screening method to predict Chlamydia trachomatis infection in women?

We evaluated the use of the leukocyte esterase test (LET) on first-catch urine specimens from women as a screening test to predict infection with Chlamydia trachomatis. For diagnosis, we used Abbott's ligase chain reaction (LCR) on urine specimens and isolation by tissue culture (TC) on cervical brushes. Of 4,053 women attending sexually transmitted disease and family planning clinics, 4.3% (n = 174) were positive by TC and 5.9% (n = 239) were positive by LCR. When LET was compared to TC, the sensitivity, specificity, positive predictive value, and negative predictive value were 54.0, 67.0, 6.8, and 97.0%, respectively. The corresponding performance of LET versus LCR was 53.1, 67.3, 10.1, and 95.8%. Almost half of the laboratory-confirmed chlamydial infections were negative by LET. The low specificity probably reflects multiple causes of pyuria in women and results in a low positive predictive value. LET is neither sensitive nor specific as a predictor of chlamydial infection and cannot be recommended for use as a screening test for C. trachomatis with first-catch urine samples from females from low- or moderate-prevalence populations.     

Prednisolone therapy of idiopathic feline lower urinary tract disease: a double-blind clinical study

A double-blind clinical study was performed to evaluate prednisolone as treatment for idiopathic feline lower urinary tract disease. No differences in response were observed in prednisolone- and placebo-treated cats.     

The significance of spinal cord compression as the initial manifestation of lymphoma

Sex hormones have profound effects on immune responses and may influence the outcome of autoimmune diseases such as rheumatoid arthritis (RA). We investigated the effect of gonadal steroids on the production of interleukin-1 (IL-1) and IL-6, cytokines believed to be important in the pathogenesis of RA. Peripheral blood mononuclear cells (PBMCs) were isolated from healthy male donors and male patients with RA, and were stimulated with lipopolysaccharide (LPS) in the presence of different concentrations of 17-beta-estradiol, progesterone or testosterone. In studies of cells from normal male donors, 17-beta-estradiol at pharmacological concentrations (> or = 10(-6) M) enhanced IL-1 and IL-6 secretion as well as the production of cell-associated IL-1. Progesterone and testosterone at similar concentrations inhibited IL-1 secretion but had no significant effect on IL-6 secretion or on the production of cell-associated IL-1. In studies of male RA donors, 17-beta-estradiol failed to enhance IL-1 or IL-6 secretion and progesterone failed to inhibit IL-1 secretion. The inhibitory effects of testosterone, however, appeared to be similar to that in normal donors. It is suggested that 17-beta-estradiol may promote IL-1 and IL-6 production and release, while gestation hormone, progesterone, and testosterone may inhibit IL-1 release in vivo. These data may partly explain the gender and age differences in the incidence of RA and the development of the disease in men with low and androgen levels.     

In vivo occupancy of the striatal dopamine uptake complex by various inhibitors does not predict their effects on locomotion

We have recently found that allogenic bone marrow transplantation (BMT) can be used to treat lupus nephritis in NZB x NZW F1 (B/W F1) and BXSB mice. To elucidate why and how glomerular damage is repaired, serial renal biopsies were carried out using B/W F1 mice before, and after BMT. Donor-derived B cells and macrophages with normal functions developed two weeks after BMT, whereas donor-derived functional T cells were generated after seven weeks of BMT. Visceral epithelial cells as well as macrophages in the glomeruli were activated (probably by T cell-derived lymphokines) at this time; they showed marked phagocytic activity, resulting in clearance of immune complexes (ICs) and repair of damaged basement membranes. These results suggest that normal T cell functions, which have the capacity to activate macrophages and epithelial cells, are essential in repairing IC-mediated glomerular damage.     

Pancreatic function and congenital duodenal anomalies

Six children operated on for congenital anomalies of the duodenum were investigated to find out if pancreatic dysfunction was associated with the duodenal malformation, even in the absence of clinical evidence of pancreatic insufficiency. None of the children had diarrhea and none requested nutritional support. Pancreatic function was assessed by enzyme activities (lipase, trypsin, and chymotrypsin) bicarbonate and calcium measurements in pancreatic juice obtained through a nasoduodenal tube under stimulation by secretin and cerulein. Results showed no significant modification in hydro-electrolytic secretion, but impairment of enzymatic secretion was seen. The physiopathological relationship between duodenal anomalies and pancreatic dysfunction is discussed.     

TGF-beta mediated G1 arrest in a human melanoma cell line lacking p15INK4B: evidence for cooperation between p21Cip1/WAF1 and p27Kip1

We have studied TGF-beta mediated G1 arrest in WM35, an early stage human melanoma cell line. These cells have lost p15INK4B expression through loss of one chromosome 9 and rearrangement of the other. In asynchronously growing WM35, TGF-beta caused reductions in cyclin D1, cyclin A and cdk4 proteins and their associated kinase activities and an increase in both p21Cip1/WAF1 and p27Kip1. These findings were confirmed in cells released from quiescence in the presence of TGF-beta, in which TGF-beta inhibited or delayed the reduction in the cdk inhibitors that normally occurs in late G1. In contrast to observations in other cell types, there was an increased association of both p21Cip1/WAF1 and p27Kip1 with cyclin D1/cdk4 and with cyclin E/cdk2 during TGF-beta mediated arrest of asynchronously growing cells. Upregulation of p21Cip1/WAF1 preceded that of p27Kip1. Furthermore, p21Cip1/WAF1 and p27Kip1 were not present in the same cdk complexes but bound distinct populations of target cdk molecules. Both p21Cip1/WAF1 and p27Kip1 immunoprecipitates from asynchronously growing cells contained active kinase complexes. These KIP-associated kinase activities were reduced in TGF-beta arrested cells. It has been proposed that in TGF-beta arrested epithelial cells, up-regulation of p15INK4B and of p15INK4B binding to cdk4 serves to destabilize the association of p27Kip1 with cyclin D1/cdk4, promoting p27Kip1 binding and inhibition of cyclin E/cdk2. Our findings demonstrate that this is not a universal mechanism of G1 arrest by TGF-beta. In TGF-beta arrested WM35, which lack p15INK4B, the increased p21Cip1/WAF1 may serve a similar function to that of p15INK4B: initiating kinase inhibition and providing an additional mechanism to supplement the effect of p27Kip1 on G1 cyclin/cdks.