Impaired endothelial antithrombotic activity following short-term interruption of continuous subcutaneous insulin infusion in type 1 diabetic patients

Review of literature has shown an increased rate of thrombotic complications in diabetic patients with frequent episodes of hyperketonemia. However, the mechanisms by which ketosis promotes vascular disease in diabetic patients are unclear. It was the aim of this study to investigate early changes in haemostatic parameters and oxidative stress markers during the hyperketonemic status which follows the interruption of continuous subcutaneous insulin infusion (CSII) in type I diabetic patients. Eight CSII-treated type I diabetic patients underwent a 4-hour pump arrest. Blood glucose, insulin and 3-hydroxybutirate were measured to verify the metabolic response. A vein-occlusive (VO) test was performed for the determination of tPA and PAI-1 activities and their antigen levels before and after the CSII arrest. Coagulation factor VII and VIII were evaluated by one-stage PT and PTT method, respectively. TF, vWF, tPA and PAI-1 antigens were determined by ELISA, whereas tPA and PAI-1 activities using chromogenic methods. Plasma malondialdehyde (MDA) and protein carbonyl groups (PCG) levels were determined by HPLC and spectrophotometry, respectively. After the insulin deprivation phase, post-VO tPA antigen level significantly decreased (P = 0.0391), whereas TF and post-VO PAI-1 activity and antigen levels significantly increased (P = 0.0156 and P = 0.0234, respectively). Plasma MDA and PCG levels were 1.88-fold and 1.74-fold higher than baseline values, respectively. In conclusion, the impairment of the fibrinolytic potential and the increases in TF, MDA and PCG levels may enhance the risk of both arterial and venous thrombosis during ketosis. Thus, early detection of hyperketonemia in DM patients could contribute to the prevention of life-threatening vascular events.     

Amyotrophic lateral sclerosis and prolactinoma

A 59-year-old male with amyotrophic lateral sclerosis is presented. Investigations revealed the co-existence of a pituitary adenoma of the prolactinoma type. This combination has not been reported before. The possible relation between endocrinological disturbances and this neurological disease is discussed.     

The Endovascular Management of Iliac Artery Aneurysms

Background: Isolated aneurysms of the iliac arteries are uncommon. Previously treated by conventional surgery, there is increasing use of endografts to treat these lesions. Purpose: The purpose of this study was to assess the efficacy, safety, and durability of the stent-grafts for treatment of iliac artery aneurysms (IAAs). The results of endografting for isolated IAAs over a 10-year period were analyzed retrospectively. The treatment methods differed depending on the anatomic location of the aneurysms. Twenty-one patients (1 woman, 20 men) underwent endovascular stent-graft repair, with one procedure carried out under emergency conditions after acute rupture. The mean aneurysm diameter was 4.6 cm. Results: The procedural technical success was 100%. There was zero 30-day mortality. Follow-up was by interval CT scans. At a mean follow-up of 51.2 months, the stent-graft patency rate was 100%. Reintervention was performed in four patients (19%): one patient (4.7%) with a type I endoleak and three patients (14.3%) with type II endoleaks. Conclusion: We conclude that endovascular repair of isolated IAAs is a safe, minimally invasive technique with low morbidity rates. Follow-up results up to 10 years suggest that this approach is durable and should be regarded as a first treatment option for appropriate candidates.     

In situ cross-linkable hyaluronan hydrogels containing polymeric nanoparticles for preventing postsurgical adhesions

OBJECTIVE: To develop a combined barrier method and drug delivery system ("hybrid system") for preventing postoperative peritoneal adhesions, which could combine the biocompatibility and ease of application of in situ cross-linkable hydrogels with the controlled release features of polymeric nanoparticles. METHODS: Poly(lactic-co-glycolic acid) nanoparticles were dispersed in aldehyde- and hydrazide-modified hyaluronic acids (HA), then combined via a double-barreled syringe. The material was subjected to mechanical testing and was assayed for in vitro cytotoxicity to murine mesothelial cells. Subsequently, it was tested for biocompatibility by intraperitoneal injection in mice. The hybrid's effectiveness in preventing postsurgical adhesions was assessed using a rabbit sidewall defect-cecum abrasion model, where it was applied to both injured surfaces. RESULTS: The in situ hybrid gel system formed a flexible and durable hydrogel in less than 10 seconds. It had low in vitro cytotoxicity. In the mouse, the cross-linked HA maintained the polymeric nanoparticles in the peritoneum for 1 week, which we had previously shown would have cleared in less than 2 days, and no animals developed adhesions. Notably, the hybrid gel, even in the absence of encapsulated drug, was highly effective in preventing peritoneal adhesions in the rabbit model employed. Animals treated with the hybrid (n = 8) had no adhesions in 62.5% of cases, and none had adhesions that could only be separated by sharp dissection. In contrast, only 4.2% of untreated animals (n = 24) had no adhesions, and 58.3% developed adhesions requiring sharp dissection. CONCLUSIONS: The hybrid cross-linked HA-nanoparticle system described here appears to be a biocompatible and highly effective adhesion barrier, which could also deliver antiadhesion drugs.     

Use of Boronic Acid Disk Tests To Detect Extended- Spectrum β-Lactamases in Clinical Isolates of KPC Carbapenemase-Possessing Enterobacteriaceae

We evaluated boronic acid (BA)-based methods for their ability to detect extended-spectrum β-lactamases (ESBLs) among clinical isolates of KPC-producing members of the Enterobacteriaceae family. A total of 155 isolates of Klebsiella pneumoniae (n = 141), Escherichia coli (n = 6), Enterobacter aerogenes (n = 6), and Klebsiella oxytoca (n = 2) genotypically confirmed to be KPC producers were analyzed. As many as 118 isolates harbored ESBLs (103 harbored SHV-type ESBLs, 13 harbored CTX-M-type ESBLs, and 2 harbored both SHV- and CTX-M-type ESBLs); the remaining 37 isolates were genotypically negative for ESBL production. The CLSI ESBL confirmatory test was positive for 79 of the 118 ESBL producers (sensitivity, 66.9%), while all 37 non-ESBL producers were negative (specificity, 100%). When a ≥5-mm increase in the zone diameter of either the cefotaxime (CTX)-clavulanate (CA) or the ceftazidime (CAZ)-CA disks containing BA compared with the zone diameter of the CTX or CAZ disks containing BA was considered to be a positive result for ESBL production, the method detected all 118 ESBL producers (sensitivity, 100%) and showed no false-positive results for non-ESBL producers (specificity, 100%). Double-disk synergy tests, in which disks of CTX, CAZ, aztreonam, or cefepime in combination with BA were placed at distances of 20, 25, and 30 mm (center to center) from a disk containing amoxicillin (amoxicilline)-clavulanate-BA, were able to detect 116 (98.3%), 101 (85.6%), and 28 (23.7%) of the ESBL-positive isolates, respectively; no false-positive results for non-ESBL-producing isolates were detected. Our results demonstrate that the modified CLSI ESBL confirmatory test with antibiotic disks containing BA is the most accurate phenotypic method for the detection of ESBLs in Enterobacteriaceae producing KPC carbapenemases.During the last decade, carbapenem resistance has emerged among clinical isolates of the Enterobacteriaceae family, and this is increasingly attributed to the production of β-lactamases capable of hydrolyzing carbapenems (23). Among these enzymes, a new type of Ambler class A β-lactamase, the Klebsiella pneumoniae carbapenemase (KPC), has been rapidly spreading among K. pneumoniae isolates and other Enterobacteriaceae in the northeastern regions of the United States and has now spread to several regions of North and South America, as well as in Israel, China, and Greece (2, 13, 16, 21).The current spread of KPC enzymes makes them a potential threat to currently available antibiotic-based treatments. These enzymes confer various levels of resistance to all β-lactams, including carbapenems, even though cefamycins and ceftazidime are only weakly hydrolyzed (15, 18). KPC-possessing strains frequently carry extended-spectrum β-lactamase (ESBL) genes (1, 3, 8, 13, 24), which could possibly contribute to the expression and dissemination of the β-lactam resistance trait (8, 18, 21). It should be also noted that KPCs and ESBLs are mostly plasmid-encoded determinants that can easily disseminate to other enterobacterial strains (3, 7, 15, 18, 26). Therefore, the phenotypic detection of ESBLs in KPC-producing isolates of the Enterobacteriaceae is of potential interest for epidemiological purposes as well as for limiting the spread of the underlying resistance mechanisms.The CLSI recommends a phenotypic confirmatory test for ESBL production that consists of measuring the growth-inhibitory zones around both cefotaxime (CTX) and ceftazidime (CAZ) disks with or without clavulanate (CA) for K. pneumoniae, Klebsiella oxytoca, Escherichia coli, and Proteus mirabilis (4). Different double-disk synergy tests (DDSTs) based on the synergy of amoxicillin (amoxicilline)-clavulanate (AMC) with extended-spectrum cephalosporins and aztreonam have also been extensively used for the detection of ESBLs (7). However, strategies for the laboratory identification of ESBLs need to be reviewed and adjusted as additional mechanisms of resistance to β-lactams coexist in enterobacterial strains (7). KPCs hydrolyze several β-lactam antibiotics, and hence, the presence of an ESBL can be masked by the expression of a KPC. Moreover, the weak inhibition of KPCs by the β-lactam inhibitors (15, 18, 30) may interfere with the interpretation of ESBL detection methods and KPC enzymes may be mistaken for ESBLs. Thus, there is a need to accurately detect ESBLs in the presence of coexisting KPC expression.Boronic acid (BA) compounds were recently reported to be reversible inhibitors of KPCs (6, 16, 27). In particular, we have shown that BA disk assays are considered positive for the detection of the KPC enzyme when the growth-inhibitory zone diameter around a meropenem, imipenem, or cefepime disk with phenylboronic acid is 5 mm or greater of the growth-inhibitory zone diameter around the disk containing meropenem or cefepime alone (27). The results of this study also showed that BA affected the activity of CAZ in ESBL-negative KPC-producing isolates but not in SHV ESBL-positive KPC-producing isolates, most likely due to the presence of the SHV ESBL, which is not restrained by BA (27). BA-based tests with disks of CAZ and CTX have also been successfully employed for the identification of ESBLs in AmpC producers (11, 25). These observations led us to design a modified CLSI ESBL confirmatory test using antibiotic disks containing BA as well as different DDSTs employing BA for the accurate detection of ESBLs in KPC-producing enterobacterial isolates.     

The m.3244G>A mutation in mtDNA is another cause of progressive external ophthalmoplegia

We sequenced all mitochondrial tRNA genes in a 61-year-old man with chronic progressive external ophthalmoplegia and mitochondrial myopathy but without mtDNA rearrangements, and identified a heteroplasmic m.3244G>A mutation in the tRNA^Leu(UUR) gene. This mutation had been previously associated with the MELAS phenotype, but not described in any detail.The mutation load in muscle was 84% and COX-negative fibers harbored greater levels of mutant genomes than COX-positive fibers. The m.3244G>A mutation affects a highly conserved nucleotide in the dihydrouridine loop and has been associated with a wobble modification deficiency of the mutant tRNA.     

The proteasome inhibitor bortezomib drastically affects inflammation and bone disease in adjuvant‐induced arthritis in rats

Objective

To explore the effect of bortezomib in splenocytes and fibroblast‐like synoviocytes (FLS) and its in vivo potency in a rat model of adjuvant‐induced arthritis (AIA), which resembles human rheumatoid arthritis (RA).

Methods

AIA was induced with Freund's complete adjuvant. Splenocyte and FLS proliferation and apoptosis were measured by radioactivity incorporation and flow cytometry, respectively. The invasiveness of FLS from rats with AIA was tested in a Transwell system. The pattern of cytokine secretion was evaluated by cytometric bead array in splenocyte supernatants. Bortezomib was administered prophylactically or therapeutically, and arthritis was assessed clinically and histologically. Immunohistochemistry was performed for markers of inflammation and angiogenesis in joints. Hematologic and biochemical parameters were tested in peripheral blood (PB). Representative animals were examined by computed tomography (CT) scanning before and after bortezomib administration. The expression of Toll‐like receptor 2 (TLR‐2), TLR‐3, and TLR‐4 in PB and FLS was measured by real‐time polymerase chain reaction, and alterations in specific cell populations in PB and spleen were determined by flow cytometry.

Results

In vitro, bortezomib exhibited significant inhibitory and proapoptotic activity in splenocytes and FLS from rats with AIA, altered the inflammatory cytokine pattern, and reduced the invasiveness of FLS from rats with AIA. In vivo, bortezomib significantly ameliorated disease severity. Remission was associated with improved histology and decreased expression of CD3, CD79a, CD11b, cyclooxygenase 1, and factor VIII in target tissues as well as down‐regulation of TLR expression in PB and cultured FLS. CT scanning demonstrated a bone healing effect after treatment.

Conclusion

Our findings suggest that bortezomib affects AIA in a pleiotropic manner and that this drug may be effective in RA.

     

Population attributable risk of tobacco and alcohol for upper aerodigestive tract cancer

Tobacco and alcohol are major risk factors for upper aerodigestive tract (UADT) cancer and significant variation is observed in UADT cancer rates across Europe. We have estimated the proportion of UADT cancer burden explained by tobacco and alcohol and how this varies with the incidence rates across Europe, cancer sub-site, gender and age. This should help estimate the minimum residual burden of other risk factors to UADT cancer, including human papillomavirus. We analysed 1981 UADT cancer cases and 1993 controls from the ARCAGE multicentre study. We estimated the population attributable risk (PAR) of tobacco alone, alcohol alone and their joint effect. Tobacco and alcohol together explained 73% of UADT cancer burden of which nearly 29% was explained by smoking alone, less than 1% due to alcohol on its own and 44% by the joint effect of tobacco and alcohol. Tobacco and alcohol together explained a larger proportion of hypopharyngeal/laryngeal cancer (PAR=85%) than oropharyngeal (PAR=74%), esophageal (PAR=67%) and oral cancer (PAR=61%). Tobacco and alcohol together explain only about half of the total UADT cancer burden among women. Geographically, tobacco and alcohol explained a larger proportion of UADT cancer in central (PAR=84%) than southern (PAR=72%) and western Europe (PAR=67%). While the majority of the UADT cancers in Europe are due to tobacco or the joint effect of tobacco and alcohol, our results support a significant role for other risk factors in particular, for oral and oropharyngeal cancers and also for UADT cancers in southern and western Europe.     

In vitro toxic effects of metal compounds on kinetic traits and ultrastructure of rabbit spermatozoa

Metal compounds have been associated with male reproductive toxicity in vivo. The aim of the present study was to investigate the in vitro effects of 20 metal compounds using rabbit ejaculated spermatozoa as a study model for spermiotoxicity. Five of the metals tested (arsenic, cadmium, chromium, mercury and vanadium) reduced sperm motility and curvilinear velocity. Ultrastructural analyses revealed three types of damage to sperm head membranes in relation to the metal used: acrosome breakage with formation of various sized microvesicles (arsenic, cadmium, mercury and platinum); a large round hole (arsenic, cadmium and chromium), and numerous folds in the acrosome membrane (vanadium). The vanadium compound, followed by chromium and mercury compounds, determined a higher number of damaged spermatozoa. In conclusion, all the studied metal compounds, at levels higher than 1 μM, may reduce sperm kinetic characteristics and probably fertilizing capacity by triggering specific morphological damages to the head and/or by inhibiting motility.