Accumulation of Clarithromycin in Macrophages Infected With Mycobacterium avium

Clarithromycin is known to accumulate in polymorphonuclear leukocytes, but no accumulation studies with macrophages have been reported. We exposed J774 macrophages, grown for 4–6 days, to clarithromycin 3.0 μg/ml for 2 hours. The cells were separated from the extracellular fluid, and the concentration of clarithromycin was determined in an agar diffusion bioassay. The accumulation of clarithromycin was 15.8-fold greater in the cells than it was in the extracellular fluid when the test was performed with noninfected cells, and 17.3-fold greater for cells infected with Mycobacterium avium. However, the ratio was substantially lower, only 3.7 for dead macrophages, suggesting that intracellular accumulation is probably an active process. These data may clarify the nature of the activity of clarithromycin against M. avium in macrophages.     

Delayed osteotomy but not fracture healing in pediatric osteogenesis imperfecta patients receiving pamidronate.

This study evaluated factors influencing fracture (n = 197) and osteotomy (n = 200) healing in children with moderate to severe OI. Pamidronate treatment was associated with delayed healing after osteotomy, but not after fracture. The data suggest that both pamidronate and mechanical factors influence bone healing in this cohort. INTRODUCTION: Intravenous pamidronate is widely used to treat children with moderate to severe osteogenesis imperfecta (OI). However, the effect of this treatment on bone healing is not well characterized. We therefore retrospectively analyzed the healing of lower limb fractures and osteotomies in children with moderate to severe OI, both before and after the start of pamidronate treatment. MATERIALS AND METHODS: Bone healing was evaluated on standard radiographs after 197 lower limb fractures (132 femur and 65 tibia) in 82 patients (age at fracture, 0.0-19.9 years) and 200 intramedullary rodding procedures in 79 patients (age at surgery, 1.2-19.8 years). Delayed healing was diagnosed when a fracture or osteotomy line was at least partially visible 12 months after the event. RESULTS: Delayed fracture healing was observed more frequently during than before pamidronate treatment. However, the effect of pamidronate was no longer significant when age differences were taken into account (odds ratio [OR], 1.76; 95% CI, 0.61-5.10). Better mobility status was a strong independent predictor of delayed healing after fractures that occurred during pamidronate treatment. After osteotomies, delayed healing was more frequent when pamidronate had been started before surgery (OR, 7.29; 95% CI, 2.62-20.3), and this effect persisted after adjustment for multiple confounders. During pamidronate treatment, older age (OR per year of age, 1.25; 95% CI, 1.06-1.47) and osteotomy of the tibia (OR, 3.51; 95% CI, 1.57-7.82) were independent predictors of delayed healing. CONCLUSIONS: This study suggests that pamidronate therapy is associated with delayed healing of osteotomy sites after intramedullary rodding procedures. Better mobility status, but not pamidronate treatment, seems to be predictive of delayed healing after fractures.     

Absence of insulin receptor gene mutations in three insulin-resistant women with the polycystic ovary syndrome

Women with polycystic ovary syndrome (PCOS) are markedly insulin-resistant, but the molecular mechanisms of these changes and their relationship to the hyperandrogenic state remain to be clarified. Mutations have recently been identified in the insulin receptor gene of patients with extreme forms of insulin resistance associated with hyperandrogenism (eg, type A insulin resistance), and these mutations account for the insulin resistance in such patients. We performed this study to determine whether mutations in the coding portion of the insulin receptor gene were responsible for insulin resistance in PCOS. Insulin binding studies using cultured skin fibroblasts of three obese (body mass index > 27 kg/m²) women with PCOS (ie, mild hyperandrogenemia and chronic anovulation of unknown etiology) and documented insulin resistance showed no apprarent abnormalities in either the number or affinity of insulin binding sites. Direct sequencing of all 22 exons of the insulin receptor gene from two of the women with PCOS did not reveal any mutations. Furthermore, both alleles of the gene were expressed at equal levels. In a third insulin-resistant PCOS woman, there was no evidence for a mutation in the coding portion of the insulin receptor gene as determined by denaturing gradient gel electrophoresis (DGGE). We conclude that the insulin resistance in these PCOS women was caused by a defect extrinsic to the insulin receptor.     

Flickering fusion pores comparable with initial exocytotic pores occur in protein-free phospholipid bilayers

For the act of membrane fusion, there are two competing, mutually exclusive molecular models that differ in the structure of the initial pore, the pathway for ionic continuity between formerly separated volumes. Because biological “fusion pores” can be as small as ionic channels or gap junctions, one model posits a proteinaceous initial fusion pore. Because biological fusion pore conductance varies widely, another model proposes a lipidic initial pore. We have found pore opening and flickering during the fusion of protein-free phospholipid vesicles with planar phospholipid bilayers. Fusion pore formation appears to follow the coalescence of contacting monolayers to create a zone of hemifusion where continuity between the two adherent membranes is lipidic, but not aqueous. Hypotonic stress, causing tension in the vesicle membrane, promotes complete fusion. Pores closed soon after opening (flickering), and the distribution of fusion pore conductance appears similar to the distribution of initial fusion pores in biological fusion. Because small flickering pores can form in the absence of protein, the existence of small pores in biological fusion cannot be an argument in support of models based on proteinaceous pores. Rather, these results support the model of a lipidic fusion pore developing within a hemifused contact site.     

The effect of growth hormone on the development of diabetic kidney disease in rats.

BACKGROUND: Nephropathy is the most severe complication of diabetes mellitus. We investigated the effect of exogenous growth hormone (GH) administration on renal function and matrix deposition in the streptozotocin (STZ) model of type I-diabetic rat. METHODS: Adult female STZ-diabetic rats (D), non-diabetic control rats injected with saline (C) and control and diabetic rats injected with bovine GH for 3 months (CGH and DGH, respectively) were used. RESULTS: The usual renal hypertrophy seen in D animals was more pronounced in the DGH group. Creatinine clearance increased only in the D rats, but not in the other groups, including DGH. Albuminuria was observed in the D animals but was significantly elevated in the DGH group. Glomeruli from DGH animals showed more extensive matrix accumulation (manifested as an increase in mesangial/glomerular area ratio). Renal extractable insulin-like growth factor (IGF-I) mRNA was decreased in the D and DGH groups, but renal IGF-I protein was not significantly increased. Renal IGF binding protein-1 was increased in the D groups and further increased in the DGH group, at both the mRNA and protein levels. CONCLUSIONS: GH-treated diabetic rats had less hyperfiltration and more albuminuria, concomitant with more glomerular matrix deposition, when compared with regular diabetic animals. This was associated with a significant increase in renal IGFBP-1, and dissociated from IGF-I changes. Thus, in this model, GH exacerbates the course of diabetic kidney disease.     

Iron-mobilizing properties of the gadolinium-DTPA complex: clinical and experimental observations.

BACKGROUND: Gadolinium (Gd) magnetic resonance imaging (MRI) contrast agents are considered to be safe in patients with impaired renal function. Our study investigates a mechanism of severe iron intoxication with life-threatening serum iron levels in a haemodialysis patient following MRI with Gd-diethylenetriaminepentaacetic acid (Gd-DTPA) administration. His previous history was remarkable for multiple blood transfusions and biochemical evidence of iron overload. We hypothesized that Gd-DTPA may have an iron-mobilizing effect in specific conditions of iron overload combined with prolonged exposure to the agent. METHODS: For the in vitro study, Gd-DTPA was added to mice liver homogenate and iron metabolism parameters were measured after incubation in comparison with the same samples incubated with saline only. For the in vivo study, an experimental model of acute renal failure in iron-overloaded rats was designed. Previously iron-overloaded and normally fed rats underwent bilateral nephrectomy by renal pedicle ligation, followed by Gd-DTPA or saline injection. Iron and iron saturation levels were checked before and 24 h after Gd-DTPA or vehicle administration. RESULTS: Significant mobilization of iron from mice liver tissue homogenate in mixtures with Gd in vitro was seen in the control (saline) and in the experimental (Gd) groups (513+/-99.1 vs 1117.8+/-360.8 microg/dl, respectively; P<0.05). Administration of Gd-DTPA to iron-overloaded rats after renal pedicle ligation caused marked elevation of serum iron from baseline 143+/-3.4 to 570+/-8 microg/dl (P<0.0001). There were no changes of the named parameter, either in iron-overloaded anuric rats after saline injection or in normal diet uraemic animals, following Gd-DTPA administration. CONCLUSIONS: The combination of iron overload and lack of adequate clearance of Gd chelates may cause massive liberation of iron with dangerous elevation of free serum iron. It is highly recommended that after Gd contrast study, end-stage renal disease patients with probable iron overload should undergo prompt and intensive haemodialysis for prevention of this serious complication.     

Numerical method for weights adjustment in minimax multi‐model LQ‐control

The minimax linear quadratic problem, where ‘max’ is taken over a finite set of indices (models) and ‘min’ is taken over the set of admissible controls, is considered. The solution is obtained by the robust optimal control application. The control turns out to be a linear combination of the controls optimal for each individual model. This paper develops a numerical method for the optimal weights adjustment. An example shows a quick convergence of the proposed procedure. Copyright © 2007 John Wiley & Sons, Ltd.     

Method for three-dimensional visualization of neurodegeneration in cupric-silver stained serial rat brain slices

The spatial distribution of neurodegeneration in brains is difficult to visualize when working from 2-D serial slices. In studies where repetitive operant behavior measurements are made over several weeks following organic solvent exposure, definitive evidence of degeneration in brain structures may have been significantly cleared by the time the tissue is prepared histologically. The only remaining evidence that injury has occurred may be nothing more than neuronal and cellular debris. By choosing stains that are specific for this type of residual and/or indicative of specific pathology, a 3-D representation of the spatial distribution of the neuronal and cellular debris fields within the organ can be highlighted and displayed. We present a method for visualizing the spatial distribution of neuronal degeneration that can result from low-level organic solvent exposure scenarios. A cupric-silver stain highly specific for neuronal degeneration is used to identify neuronal debris fields in 73 serial slices of brains of rodents that were exposed to toluene vapors. Serial brain sections stained with cupric-silver are scanned at 600 dpi using a gray-scale protocol. Using commercially available software, scans are assembled into 3-D images showing both topographical and internal anatomical details. The reassembled images are further processed into stereo pairs. Gray-scale scans are compared to the original sections to establish gray-scale ranges for healthy and damaged tissue and artifact staining.     

MICA4/HLA-DRB1*04/TNF1 haplotype is associated with mixed connective tissue disease in Swedish patients

In order to investigate major histocompatibility complex (MHC) class I chain-related gene A (MICA), tumor necrosis factor (TNFa), -308TNFA, and human leukocyte antigen (HLA-DR/DQ) polymorphisms in mixed connective tissue disease (MCTD), we analyzed 24 patients and 229 healthy controls from Sweden. MICA and TNFa typing was performed by polymerase chain reaction (PCR) and genotyping. HLA-DR and -DQ were genotyped using PCR-sequence specific primers (PCR-SSP) and PCR-sequence-specific oligonucleotide probe (PCR-SSOP), respectively. For analysis of -308TNFA polymorphisms we performed PCR with restriction endonuclease enzymes. We found that the MICA5.1-5.1 genotype was positively associated with MCTD. Shared epitope genes (DRB1*01 and DRB1*04) were also significantly positively associated with MCTD. Polymorphism of -308TNFA was not differently distributed in MCTD patients compared with controls. Furthermore, we demonstrated that frequencies of three estimated haplotypes were increased in MCTD patients compared with controls. Interestingly, the haplotype with MICA allele 4 together with DRB1*04 and TNF1 alleles gives the most specific pattern for MCTD patients compared with controls. Our study demonstrates a clear contribution of HLA loci in susceptibility to MCTD in the Swedish population. Susceptibility to MCTD may be linked to the MICA4/HLA-DRB1*04/TNF1 haplotype and MICA 5.1-5.1 genotype. Mixed connective tissue disease was also associated with shared epitope genes, which in RA has been associated with a more severe disease. Whether these genotypes affect the clinical phenotype of MCTD needs to be determined.     

Hydrogels in endovascular embolization. II. Clinical use of spherical particles

In this study we report the results of clinical experiments, obtained with spherical particles made from poly(2-hydroxyethyl methacrylate) used in the embolization of arteriovenous anastomoses, in the suppression of pulmonary haemorrhage and haemoptysis and in the occlusion of some other arteries. So far we have used these particles in the treatment of 187 patients. It must be stressed that the advantage of spherical particles consists in the simplicity of their introduction into the blood vessel through a catheter, while in the blood vessel itself the particle swells in blood still more, when compared with the particle size in saline. This results in an immediate and permanent haemostatic effect. No revascularization occurs.