(13)N-ammonia PET as a measurement of hindlimb perfusion in a mouse model of peripheral artery occlusive disease.

Peripheral arterial occlusive disease (PAOD) is a leading cause of mortality and morbidity in the western world. The development of noninvasive methods for assessment and comparison of the efficacy of novel therapies in animal models is of great importance. METHODS: Hindlimb ischemia was induced in nude mice by ligation and excision of the left femoral artery (n = 5) or the left iliac artery (n = 10). Assessment of limb perfusion was performed by small-animal PET analysis after intravenous injection of (13)N-ammonia between 24 h and 30 d after surgery using the ratio of perfusion between the left limb (ischemic) and the right limb (control). Activity concentration per area unit was calculated in regions of interest placed on 1-mm-thick images for numeric calculations, and the iliac and the femoral models were compared. In addition, histopathologic studies were performed to assess the degree of necrosis (hematoxylin-eosin) and fibrosis (sirius red). Immunohistochemistry analyses for identification of arterioles (alpha-smooth muscle actin) and endothelium-capillaries-(Bandeiraea simplicifolia I [BS-I] lectin) were also performed. RESULTS: Perfusion in both hindlimbs of control animals was similar (median of the left-to-right ratio = 0.99). Twenty-four hours after ischemia, perfusion of the ischemic limb (% mean +/- SD) was 33.3 +/- 10.6 and 22.1 +/- 9.9 in the femoral and iliac models, respectively. Spontaneous recovery of perfusion in the hindlimb that underwent surgery was significantly lower in the iliac model at day +15 (73.2 +/- 15.5 vs. 51.9 +/- 11.3; P < 0.01). Fibrosis increased progressively until day +30, whereas muscle necrosis was maximal at day +7 with a moderate reduction by day +30. In accordance with this positive effect, there was a statistically significant increase in the area covered with smooth muscle-coated vessels (arterioles) at day +30 in comparison with day 7 (P < 0.05). In addition, a correlation between (13)N-ammonia uptake and the amount of necrosis (r = -0.73; P = 0.06) and fibrosis (r = -0.67; P = 0.05) at day +30 was found. CONCLUSION: (13)N-Ammonia imaging allows semiquantitative evaluation of hindlimb perfusion in surgical mouse models of acute hindlimb ischemia. Although spontaneous perfusion recovery is observed in both models, the iliac model shows a substantially lower recovery and is hence better suited for assessment of new therapeutic strategies for acute hindlimb ischemic disease.     

High prevalence of anti-C1q antibodies in biopsy-proven active lupus nephritis.

BACKGROUND: Anti-C1q antibodies (anti-C1q) have been shown to correlate positively with systemic lupus erythematosus (SLE) nephritis. Several clinical studies indicated a high negative predictive value, suggesting that active lupus nephritis is rarely seen in patients with no anti-C1q. However, the true prevalence of anti-C1q at the time of active lupus nephritis has not been well established. The aim of this study was to determine prospectively the prevalence of anti-C1q in proven active lupus nephritis at the time of the renal biopsy. METHODS: In this prospective multi-centre study, we investigated adult SLE patients undergoing renal biopsy for suspected active lupus nephritis. Serum samples were taken at the time of the biopsy and analysed for the presence of anti-C1q in a standardized way. The activity of lupus nephritis was classified according to the renal histology. Biopsies were also analysed for the presence of glomerular IgG, C1q and C3 deposition. RESULTS: A total of 38 patients fulfilling at least 4/11 American College of Rheumatology (ACR) criteria for the diagnosis of SLE were included. Out of this, 36 patients had proliferative (class II, III or IV) and two had class V lupus nephritis. All but one patient with proliferative lupus nephritis were positive for anti-C1q (97.2%) compared with the 35% of control SLE patients with inactive lupus nephritis and 25% of SLE patients without lupus nephritis ever. All patients were positive for glomerular C1q (36/36) and 37/38 patients had glomerular IgG deposits. Anti-C1q strongly decreased during successful treatment. CONCLUSIONS: Anti-C1q have a very high prevalence in biopsy-proven active lupus nephritis, thus a negative test result almost excludes active nephritis. The data support the hypothesis of a pathogenic role of anti-C1q in lupus nephritis.     

Study of the cytotoxicity and antioxidant capacity of N/OFQ(1–13)NH2 and its structural analogues

The effect of side-chain shortening of N/OFQ(1–13)NH₂ at position 9 ([Orn⁹]N/OFQ(1–13)NH₂, [Dab⁹]N/OFQ(1–13)NH₂, [Dap⁹]N/OFQ(1–13)NH₂) was studied regarding potential toxicity and antioxidant capacity. Staurosporine- and H2O2-induced damage of SH-SY5Y neuroblastoma cells was not changed in the presence of N/OFQ(1–13)NH2 and [Orn⁹]N/OFQ(1–13)NH₂, but was strongly enhanced in the presence of [Dab⁹]N/OFQ(1–13)NH₂ and [Dap⁹]N/OFQ(1–13)NH₂. Moreover, treatment of cells with the latter two analogues alone led to cell injury. Neuropeptide-dependent differences in the viability of SH-SY5Y cells were also observed, i.e., a cytoprotective effect was observed only in the presence of N/OFQ(1–13)NH₂ and [Orn⁹]N/OFQ(1–13)NH₂. Compared to [Dab⁹]N/OFQ(1–13)NH₂ and [Dap⁹]N/OFQ(1–13)NH₂, the effects of N/OFQ(1–13)NH₂ and [Orn⁹]N/OFQ(1–13)NH₂ were more beneficial in systems generating free oxygen radicals (O2^? and OH), as well as on the antioxidant status of rat brain and liver. Taken together, our findings show that N/OFQ(1–13)NH₂ and its structural analogue [Orn⁹]N/OFQ(1–13)NH₂ possess more favorable profiles than the other two nociceptin (N/OFQ) analogues. The present results suggest that shortening of the side-chain of N/OFQ(1–13)NH2 might increase cell damage and reduce the viability of SH-SY5Y neuroblastoma cells. Moreover, such alterations may lead to changes in free-oxygen generating systems and in antioxidant status in animal tissues.     

Time-dependent changes in cardiac growth after kidney transplantation: the impact of pre-dialysis ventricular mass.

BACKGROUND: Left ventricular hypertrophy (LVH) is common in chronic kidney disease (CKD), including kidney transplant recipients. However, time-related left ventricular mass changes (DeltaLVM) from pre-dialysis stage to beyond the first post-transplant year have not been clearly identified. METHODS: We studied a cohort of 60 stages 4-5 CKD patients without overt cardiac disease, who underwent three echocardiograms during follow-up: at pre-dialysis stage, on dialysis and after kidney transplantation (KT). Multiple linear regression was used to model DeltaLVM from baseline study. Cox proportional analysis was used to determine risk factors associated with either de novo LVH or>20% DeltaLVMI over time. RESULTS: Patients with baseline LVH (n=37; 61%) had a higher body mass index (BMI) than those without LVH (n=23; 39%) (P=0.013). BMI, haemoglobin levels (P=0.047) and non-use of angiotensin-converting enzyme inhibitors (ACEI) (P=0.057) were associated with baseline left ventricular mass index (LVMI). Twelve out of 23 patients (52%) with normal LVM at baseline, developed either de novo LVH or>20% DeltaLVMI at follow-up. On the other hand, 29 (78%) of those with initial LVH maintained this abnormality, and 8 (22%) normalized LVM post-transplantation. Factors associated with DeltaLVMI were age (P=0.01), pre-dialysis LVMI (P<0.0001), serum creatinine (P=0.012) and the use of ACEI post-transplantation (P=0.009). In Cox analysis, pre-dialysis LVMI was associated with de novo LVH or>20% DeltaLVMI over time (hazard ratio 1.009; 95% confidence interval 1.004 to 1.015; P=0.001). CONCLUSIONS: Successful KT may not completely normalize LVM post-transplantation. Pre-dialysis LVMI, traditional risk factors and no use of ACEI may perpetuate cardiac growth following KT.     

Impact of cold ischemia time on renal allograft outcome using kidneys from young donors

Prolonged cold ischemia time (CIT) is associated with delayed graft function and worse kidney transplant (KT) outcome, but the effect of CIT on long‐term allograft survival in KT from younger donors has not been well established. We investigated the predictive value of CIT exposure on long‐term death‐censored graft loss in 829 KT recipients from younger donors (<50 years) that were performed in our center between 1991 and 2005. Overall death‐censored graft failure rate was significantly higher in CIT≥19 h group versus CIT<19 h group (26 vs. 16.5%; P = 0.002). Significant differences were also observed when patients with primary nonfunctioning graft were excluded (21 vs. 14%; P = 0.020) and in patients who received tacrolimus plus mycophenolate mofetil (12 vs. 4%; P = 0.05). By multivariate Cox analysis, CIT was found to be independently associated with death‐censored graft loss with a 20% increase for every 5 h of CIT [relative risk (RR) 1.04; 95% Confidence Interval (CI): 1.01–1.1; P = 0.021]. Likewise, graft loss risk significantly increased in CIT≥19 h group versus CIT<19 h group (RR 1.5; 95%CI: 1.1–2.1; P = 0.023). Prolonged CIT is an independent predictor of graft survival in KT from younger donors. Efforts at minimizing CIT (<19 h) should improve transplant outcome significantly in this population.     

Dietary Doses of Sulforaphane Affect Hepatic Drug Metabolizing Enzymes in Spontaneously Hypertensive Rats

We previously demonstrated that exposure of spontaneously hypertensive rats (SHR) to dietary doses of sulforaphane (SF) results in resisting the progressive rise in blood pressure that is normally seen in these rats. This study investigates the potential effect of SF on hepatic drug metabolizing enzymes (DME) in SHR. The activity and/or protein expression of selected cytochrome P450 (CYP) enzymes and microsomal epoxide hydrolase (mEH) were measured in hepatic microsomes using specific probe substrates and/or polyclonal antibodies. Cytosolic fraction was utilized to measure protein level and activity of major antioxidant systems. The high dose SF resulted in a significant reduction of activity and apoproteins level of CYP1A2 and CYP2C9 and activities of CYP2B1/2 and mEH. No effect of SF was observed on the rest of the studied CYP enzymes. Both doses of SF resulted in a significant induction of both hepatic glutathione level and activities of superoxide dismutase and catalase. Activities of hepatic glutathione‐S‐transferases, glutathione reductase, and glutathione peroxidase were significantly induced only with the high dose. This study demonstrates that dietary doses of SF modulate the activity or protein expression of DME. Additionally, induction of the impaired antioxidant system in SHR may explain the blood pressure lowering effect of SF in this rat model. Copyright © 2015 John Wiley & Sons, Ltd.     

Topical fluorometholone treatment and desiccating stress change inflammatory protein expression in tears

Purpose

It was hypothesized that tear protein biomarkers could predict the effects of topical steroid treatment and desiccating stress in patients with dry eye disease (DED). To test this concept, a randomized, double-masked, controlled clinical trial with 41 patients was conducted.

A new CACNA1A gene mutation in acetazolamide-responsive familial hemiplegic migraine and ataxia.

OBJECTIVE: To search for mutations in the calcium channel gene CACNA1A and to study the genotype-phenotype correlation in a family with a severe familial hemiplegic migraine (FHM) phenotype and a slowly progressive cerebellar ataxia. BACKGROUND: CACNA1A gene mutations on chromosome 19 are involved in approximately 50% of FHM families. The association of FHM and cerebellar ataxia has been reported in a small number of FHM families, all linked to chromosome 19. METHODS: The proband, in addition to typical hemiplegic migraine attacks, experienced severe episodes during which hemiplegia was associated with acutely altered consciousness and fever lasting several days. She, as well as her affected sister, developed a permanent, late-onset cerebellar ataxia and cerebellar atrophy evident on MRI. Linkage analysis was performed and the whole CACNA1A gene, 47 exon-intron boundaries, was analyzed by double gradient-denaturing gradient gel electrophoresis (DG-DGGE). RESULTS: Genetic studies suggested linkage to chromosome 19p13, and DG-DGGE analysis detected a heteroduplex fragment in exon 13 of the CACNA1A gene. By direct sequencing, a G-to-A substitution resulting in an arginine to glutamine change at codon 583 in the second putative voltage sensor domain of the channel alpha1A-subunit, was identified, possibly representing the disease-causing mutation. The proband and her affected sister were treated with acetazolamide, reporting freedom from new FHM attacks but no benefit in the progression of ataxia. CONCLUSIONS: The combination of episodic dysfunction and permanent deficit could depend on the variety of functions of calcium channels and their distribution in the nervous system.     

Intraoperative Radiotherapy with Balloon-Based Electronic Brachytherapy System—A Systematic Review and First Bulgarian Experience in Breast Cancer Patients

(1) Background: We aimed to analyze currently available studies with intraoperative radiotherapy (IORT) as a choice of treatment where the Xoft Axxent^® electronic brachytherapy (eBx) system was used as a single-dose irradiation and an exclusive radiotherapy approach at the time of surgery in patients with early breast cancer (EBC). We also compared the results of the systematic review to the Bulgarian experience. (2) Methods and Materials: We performed a systematic review of the studies published before February 2021, which investigate the application of a single-fraction 20 Gy radiation treatment, delivered at the time of lumpectomy in EBC patients with the Xoft Axxent^® eBx System. A systematic search in PubMed, Scopus, and ScienceDirect was performed. The results are reported following the PRISMA guidelines. The criteria on patients’ selection for IORT (the additional need for EBRT), cosmetic outcomes, and recurrence rate from the eligible studies are compared to the treatment results in Bulgarian patients. (3) Results: We searched through 1032 results to find 17 eligible studies. There are no published outcomes from randomized trials. When reported, the cosmetic outcomes in most of the studies are defined as excellent. The observed recurrence rate is low (1–5.8%). Still, the number of patients additionally referred to postoperative external breast radiotherapy (EBRT) is up to 31%. Amongst the 20 patients treated in Bulgaria, the cosmetic outcomes are also evaluated as excellent, five of which (25%) are referred for EBRT. Within median follow-up of 39 months, there was one local and one distal recurrence. (4) Conclusions: Current evidence demonstrates the Xoft Axxent^® eBx system as a safe and feasible technique for IORT delivery in EBC patients. There are no randomized controlled trials conducted at this time point to prove its long-term effectiveness. Better patient selection and a reimbursement strategy have to be proposed to extend the application of this technique in Bulgaria.