Spin states of C-60(3-) and C120On- (n=2, 3, 4) anions using electron spin transient nutation spectroscopy

Electron spin transient nutation (ESTN) experiments show that the spin multiplicity of the ground state of C-60(3-) in frozen solution is a doublet with S = 1/2. In purified samples, there is no evidence for excited states or other species with higher multiplicity. In the anions Of C120On- (n = 2, 3, 4), where the CW EPR experiments have shown that a mixture of species is present, ESTN experiments confirm that a doublet with S = 1/2 is associated with the 3- anion and triplets with S = 1 are associated with the 2- and 4- anions. A weak nutation peak attributable to m(s) = -1/2 1/2 transitions within a quartet state may arise from association of anions with spins of 1/2 and 1 in solute aggregates.

Targeting Antigen-loaded Liposomes to Myeloid Antigen Presenting Cells

Objective: To target antigen-loaded liposomes to myeloid APC in vivo for immunotherapy and to manipulate immune function through liposome composition. Method: Liposomes were loaded with ovalbumin, the lipophilic red fluorescent marker, DiI, with or without QuilA adjuvant then injected either i.v. or s.c. to naı¨ ve C57Bl/6 mice. Spleen, liver and draining LN were stained with MHC class II and various myeloid markers to determine the uptake of liposomes. Frozen sections of spleen and draining LN were stained with FITC-labeled mAb to determine which cells take up the liposomes. To determine the effect on OVA-specific T cell responses, liposomes were administered to Balb/c mice which received DO11.10 OVAspecific TCR transgenic T cells labelled with CFSE. Results: The DiI fluorescence was visualized in MHC class II+ macrophages and DC in draining lymph nodes after s.c. injection and in spleen and liver after i.v injection. Immunofluorescence microscopy shows liposome uptake in marginal zone macrophages and some DC in the T cell areas of the spleen after i.v. injection. Administration of ova-liposomes with or without QuilA stimulated a specific T cell response as measured by CFSE dilution. Conclusion: APC of liver, spleen and LN, and subsequent antigen presentation to T cells can be targeted for immunotherapy by the administration of liposomes encapsulating antigen and adjuvant. Varying the composition and routes of liposome administration is expected to alter the function of the targeted APC and the T cell response.

The systematic use of intraoperative vascular Doppler ultrasound during microsurgical subinguinal varicocelectomy improves precise identification and preservation of testicular blood supply

Objective: To evaluate the impact of systematic use of intraoperative Doppler ultrasound during microsurgical subinguinal varicocele repair. Design: Prospective clinical study. Setting: Andrology laboratory and male infertility section of the urology department of a tertiary care hospital. Patient(s): Two hundred and thirteen men with clinical varicocele. Intervention(s): Subinguinal microsurgical varicocele ligation using an intraoperative vascular Doppler flow detector. Main Outcome Measure(s): Number of veins ligated, lymphatic spared, arteries identified or accidentally ligated. Result(s): A statistically significant greater number of arteries were identified and preserved when intraoperative vascular Doppler was used. In addition, the average number of internal spermatic veins ligated was statistically significantly greater in the same group. Accidental artery ligation occurred in two cases (1.1%) in which the Doppler was not applied. There was no statistically significant difference in number of lymphatics spared between groups. Conclusion(s): Our findings showed that concomitant use of intraoperative vascular Doppler during microsurgical varicocelectomy allows more arterial branches to be preserved, and more internal spermatic veins are likely to be ligated. This device should be considered an attractive tool to improve surgical outcomes and safety. (Fertil Steril (R) 2010; 93: 2396-9. (C)2010 by American Society for Reproductive Medicine.)

Modelling of species in hood fires by conditional moment closure

Carbon monoxide, the chief killer in fires, and other species are modelled for a series of enclosure fires. The conditions emulate building fires where CO is formed in the rich, turbulent, nonpremixed flame and is transported frozen to lean mixtures by the ceiling jet which is cooled by radiation and dilution. Conditional moment closure modelling is used and computational domain minimisation criteria are developed which reduce the computational cost of this method. The predictions give good agreement for CO and other species in the lean, quenched-gas stream, holding promise that this method may provide a practical means of modelling real, three-dimensional fire situations. (c) 2005 The Combustion Institute. Published by Elsevier Inc. All rights reserved.

Impact of clinical varicocele and testis size on seminal reactive oxygen species levels in a fertile population: a prospective controlled study

Objective: To investigate: 1) the impact of clinical varicocele on reactive oxygen species (ROS) levels in neat and washed semen in a proven fertile population; and 2) the correlation between ROS levels, testicular volume, and varicocele grade in the same population of fertile men. Design: Prospective controlled clinical study. Setting: Andrology laboratory at tertiary-care hospital. Patient(s): One hundred fourteen healthy fertile men (81 normal fertile and 33 fertile with clinical varicocele) and 30 infertile patients (control subjects). Intervention(s): Standard semen analysis and measurement of sperm ROS production. Main Outcome Measure(s): Seminal parameters, seminal ROS levels, seminal leukocyte levels, clinical varicocele, and testis size. Result(s): Thirty-three of the 11.4 (29%) fertile men had clinical varicocele (grade 1, n = 14; grade 2, n = 11; and grade 3, n = 8), and the remaining 81 (71%) had a normal physical examination. Levels of ROS and semen quality did not differ significantly between the fertile men with or without varicocele. No significant differences in ROS levels in neat and washed semen were observed compared with fertile men with grades 2 and 3 varicocele and with fertile men with varicocele grade 1. The ROS levels in neat and washed semen were not significantly correlated with varicocele grade in fertile men. No significant correlations between ROS levels and testis volume were observed between the fertile groups. Conclusion(s): The presence of clinical varicocele in fertile men is not associated with higher seminal ROS levels or abnormal semen parameters. Levels of ROS are not correlated with varicocele grade or testis volume in the same population of fertile men.

Microsurgical Anatomy of the Posterior Cerebral Artery in Three-Dimensional Images

OBJECTIVE: To describe the microsurgical anatomy, branches, and anatomic relationships of the posterior cerebral artery (PCA) represented in three-dimensional images. METHODS: Seventy hemispheres of 35 brain specimens were studied. They were previously injected with red silicone and fixed in 10% formalin for at least 40 days. Four of the studied specimens were frozen at -10 degrees to -15 degrees C for 14 days, and additional dissection was done with the Klingler`s fiber dissection technique at x6 to x40 magnification. Each segment of the artery was measured and photographed to obtain three-dimensional stereoscopic images. RESULTS: The PCA origin was in the interpeduncular cistern at the pontomesencephalic junction level in 23 specimens (65.7%). The PCA was divided into four segments: P1 extends from the PCA origin to its junction with the posterior communicating artery with an average length of 7.7 mm; P2 was divided into an anterior and posterior segment. The P2A segment begins at the posterior communicating artery and ends at the most lateral aspect of the cerebral peduncle, with an average length of 23.6 mm, and the P2P segment extends from the most lateral aspect of the cerebral peduncle to the posterior edge of the lateral surface of the midbrain, with an average length of 16.4 mm; P3 extends from the posterior edge of the lateral surface of the midbrain and ends at the origin of the parieto-occipital sulcus along the calcarine fissure, with an average length of 19.8 mm; and the P4 segment corresponds to the parts of the PCA that run along or inside both the parieto-occipital sulcus and the distal part of the calcarine fissure. CONCLUSIONS: To standardize the neurosurgical practice and knowledge, surgical anatomic classifications should be used uniformly and further modified according to the neurosurgical experience gathered. The PCA classification proposed intends to correlate its anatomic segments with their required microneurosurgical approaches.

In situ hybridization detection of homeobox genes reveals distinct expression patterns in oral squamous cell carcinomas

Aims: To analyse the expression of three homeobox genes (HOXA7, PITX1 and PRRX1) in oral squanous cell carcinomas (OSCC) and the relationship of such expression to certain distinct histopathological features of OSCC and in comparison to adjacent non-neoplastic epithelium (NT). Methods and results: Digoxigenin-labelled riboprobes that are specific for each homeobox gene were generated and in situ hybridization was carried out on frozen sections. In NT samples, HOXA7 and PITX1 transcripts were found more frequently in all epithelial layers, while PRRX1 was expressed in the basal layer. With OSCC samples, expression of the three genes was associated with all histological features. However, the HOXA7 and PITX1 signals were more intense in sheets and nests and PRRX1 in small nests and isolated cells. Conclusion: HOXA7, PIXT1 and PRRX1 homeobox genes have different patterns of expression in OSCC depending on its histological features.

Analysis of Different Staining Techniques for C4d Detection in Renal Allograft Biopsies

Capillary C4d deposition has been recognized as a marker of antibody-mediated rejection (AMR). Although the detection of capillary C4d by means of immunofluorescence (IF) in cryostat sections is well established, frozen tissue is not always available, thus limiting the diagnosis of AMR. The aim of the present study was to analyze different techniques for C4d staining and the prevalence of C4d in renal allograft biopsies. Detection of C4d was carried out using IF or immunohistochemistry (IHC) on frozen and paraffin sections of renal allograft biopsies available from the same patients. Biopsies obtained from 20 patients were classified into 3 groups: no rejection, acute rejection, and chronic allograft nephropathy (CAN). The capillary C4d deposition prevalence in frozen-IF, considered the gold standard technique for C4d detection, was 45% (9/20 cases). Compared with frozen-IF, the frozen-IHC technique presented an 85% concordance rate (17/20 cases; r =.70; P <.001; sensitivity = 77.8%; specificity = 90.9%). The paraffin-IF technique showed similar results, with an 80% concordance rate (16/20 cases; r =.64; P <.005; sensitivity = 55.6%; specificity = 100%), whereas C4d detection occurred in only 65% of paraffin-IHC cases (13/20; r =.30; not significant; sensitivity = 66.7%; specificity = 63.6%). No capillary C4d deposition was detected in cases without evidence of rejection. However, 4/7 cases (57%) of acute rejection were C4d positive. In the CAN group, 5111 cases (45%) were C4d positive. In conclusion, these results demonstrated that frozen-IHC and paraffin-IF can be considered alternative techniques to frozen-IF for C4d detection. The paraffin-IHC technique displayed the lowest concordance rate for C4d detection.

Testicular Sertoli cell function in male systemic lupus erythematosus

Objective. To assess the testicular Sertoli cell function in male SLE patients. Methods. Thirty-four consecutive patients were prospectively selected to evaluate serum inhibin B. Clinical features, treatment, semen analysis, urological evaluation, testicular ultrasound, hormones and anti-sperm antibodies were determined. Results. Patients were subdivided into two groups: low serum inhibin B (Group 1, n = 8) and normal levels (Group 2, n 26). The median sperm concentration (P = 0.024), total sperm count (P = 0.023) and total motile sperm count (P = 0.025) were lower in Group 1. Inhibin B levels were positively correlated with sperm concentration (r = 0.343), total motile sperm count (r = 0.357), and negatively correlated with follicule-stimulating hormone (FSH) (r = 0.699) and luteinizing hormone (r = 0.397). The median serum inhibin B was lower in SLE patients treated with intravenous cyclophosphamide (IVCYC) compared with those without this therapy (P = 0.031). Further evaluation of the 26 SLE patients with normal inhibin B and FSH levels revealed that medians of inhibin B/FSH ratio were lower in SLE patients with oligozoospermia compared with normozoospermia (P = 0.004). This ratio was also lower in SLE patients treated with IVCYC than those without this therapy (P = 0.04). In contrast, inhibin B serum level alone did not discriminate the later group of patients (P = 0.12). Conclusions. This is the first study to identify a high frequency of testicular Sertoli cell dysfunction in male SLE associated with semen abnormalities. Further prospective studies are necessary to determine if inhibin levels and inhibin B/FSH ratio will be an earlier and useful marker of IVCYC toxicity in these patients.

High-titer antichromatin antibody is associated with proliferative class IV of lupus nephritis

The antichromatin antibody (aCT) has been described as a useful marker for lupus nephropathy. The relevance of its nephritogenic potential may be appropriately evaluated in the context of renal histopathology. Therefore, the present study investigated the relationship of aCT with a particular histopathologic class of lupus nephritis (LN). Seventy-eight consecutive patients with systemic lupus erythematosus (ACR criteria) and active nephritis who underwent renal biopsy from 1999 to 2004 and with available frozen serum sample obtained at the time of biopsy were selected. aCT was measured by ELISA, and anti-dsDNA was measured by indirect immunofluorescence (IIF) and by ELISA. All renal biopsies were revised in a blinded manner by the same expert renal pathologist. Charts were extensively reviewed for demographic and renal features obtained at the time of biopsy. The prevalence of aCT (>= 20 U) was 59% with a mean titer of 74.3 +/- 38.7U. Both aCT-positive and aCT-negative groups of patients had similar age, gender distribution, duration of lupus, and duration of renal disease. Anti-dsDNA was detected by IIF in 29.5% and by ELISA in 42.3% of the patients. Concomitant presence of both antibodies was observed in 63% (29/46) [anti-dsDNA by ELISA] and 45.6% (21/46) [anti-dsDNA by IIF] of the patients. Lower serum levels of C3 (73% vs. 40%, P=0.0058) and C4 (82% vs. 46.7%, P=0.0021) were more commonly observed in aCT >= 20 U patients compared to the aCT-negative group. It is important to note that the use of a higher cut-off value (>= 40 U) for aCT test revealed a predominance of class IV LN (58% vs. 33%, P=0.039) in aCT >= 40 U compared to aCT<40 U group. The mean levels of proteinuria, serum albumin, and creatinine were markedly altered but were comparable in both groups (P >= 0.05). One fourth (26.3%) of the 19 patients with class IV LN and aCT >= 40 U had no detectable anti-dsDNA (ELISA). These data suggest that high-titer aCT seems to be a valuable biomarker for proliferative class IV of LN.

Male gender results in more severe lupus nephritis

Gender may produce different characteristics in the manifestation of systemic lupus erythematosus (SLE). The present study investigated the influence of gender on clinical, laboratory, autoantibodies and histopathological classes of lupus nephritis (LN). As much as 81 patients diagnosed with SLE (ACR criteria) and active nephritis, who underwent renal biopsy between 1999 and 2004, and who had frozen serum samples and clinical data available from the time of biopsy, were selected for this study. The presence of anti-P and antichromatin antibodies was measured using ELISA, and anti-dsDNA was measured using indirect immunofluorescence. All of the renal biopsies were reviewed in a blinded manner by the same expert renal pathologist. The charts were extensively reviewed for demographic and renal features obtained at the time of the biopsy. Of the 81 patients (13.6%), 11 were male SLE patients. Both male and female lupus patients were of similar age and race, and had similar durations of lupus and renal disease. The female patients had more cutaneous (95.7 vs. 45.5%, P = 0.0001) and haematological (52.9 vs. 18.2%, P = 0.04) involvements than the male SLE patients. In addition, the articular data, central nervous system analyses, serositis findings and SLEDAI scores were similar in both experimental groups. Positivity for anti-dsDNA, anti-ribosomal P and antichromatin did not differ between the two groups, and both groups showed similarly low C3 or C4 serum levels. Our analysis indicated that no histopathological class of LN was predominant in both males and females. Interestingly, the serum creatinine levels were higher in the male SLE patients compared to the female SLE group (3.16 +/- A 2.49 vs. 1.99 +/- A 1.54 mg/dL, P = 0.03), with an increased frequency of high creatinine (81.8 vs. 47.1%, P = 0.04) as well as renal activity index (7.6 +/- A 3.5 vs. 4.8 +/- A 3.5, P = 0.02). In addition, whilst the mean levels of proteinuria, cylindruria and serum albumin were markedly altered, they were comparable between both lupus men and women. Moreover, the frequencies of dialysis, renal transplantation and death were similar between the two groups. These data suggest that male patients had a more severe LN compared to women diagnosed with this renal abnormality.

Organ-Sparing Microsurgical Resection of Incidental Testicular Tumors Plus Microdissection for Sperm Extraction and Cryopreservation in Azoospermic Patients: Surgical Aspects and Technical Refinements

INTRODUCTION The management of nonpalpable testicular masses is a challenging task, and coexisting infertility can further complicate the treatment decisions. We present our technique for microsurgical organ-sparing resection of incidental nonpalpable testicular nodules combined with microdissection for testicular sperm extraction and tissue cryopreservation in azoospermic patients. TECHNICAL CONSIDERATIONS Five infertile patients with azoospermia presented with nonpalpable hypoechoic testicular masses that were detected by Ultrasonography and underwent organ-sparing surgery. The testis was delivered through an inguinal incision, and the blood circulation was interrupted with a vascular clamp placed on the spermatic cord. Sludged ice was used to prevent warm ischemia, and a temperature probe was used to control the temperature at 12 degrees-15 degrees C. Real-time reflex ultrasonography was used to locate the tumor, and a stereotaxic hook-shaped needle was inserted under ultrasound guidance. The needle was placed adjacent to the tumor to guide the microsurgical resection. The tunica albuginea was incised over the tumor, which was dissected and removed, along with the adjoining parenchymal tissue. Frozen section studies were performed and, if malignancy was confirmed, biopsies of the tumor cavity margins and remaining parenchyma were obtained to ensure the absence of residual tumor. Microdissection was performed for excision of selected enlarged tubules that were processed and cryopreserved. CONCLUSIONS We present a technique for microsurgical organ-sparing resection of testicular tumor and sperm extraction that can be used in selected infertile patients with azoospermia in whom incidental masses have been diagnosed by ultrasonography. This conservative approach should be especially considered for patients with a solitary testis or bilateral tumors. UROLOGY 73: 887-892, 2009. (C) 2009 Elsevier Inc.

Magnetic characterization by SQUID and FMR of a biocompatible ferrofluid based on Fe(3)O(4)

Biocompatible superparamagnetic iron oxide nanoparticles of magnetite coated with dextran were magnetically characterized using the techniques of SQUID (superconducting quantum interference device) magnetometry and ferromagnetic resonance (FMR). The SQUID magnetometry characterization was performed by isothermal measurements under applied magnetic field using the methods of zero-field-cooling (ZFC) and field-cooling (FC). The magnetic behavior of the nanoparticles indicated their superparamagnetic nature and it was assumed that they consisted exclusively of monodomains. The transition to a blocked state was observed at the temperature T(B) = (43 +/- 1) K for frozen ferrofluid and at (52 +/- 1) K for the lyophilized ferrofluid samples. The FMR analysis showed that the derivative peak-to-peak linewidth (Delta H(PP)), gyromagnetic factor (g), number of spins (N(S)), and spin-spin relaxation time (T(2)) were strongly dependent on both temperature and super-exchange interaction. This information is important for possible nanotechnological applications, mainly those which are strongly dependent on the magnetic parameters.

A method to immunolabel rodent spinal cord neurons and glia for molecular study in specific laser microdissected cells involved in neurodegenerative disorders

Neuron-glia interaction is involved in physiological function of neurons, however, recent evidences have suggested glial cells as participants in neurotoxic and neurotrophic mechanisms of neurodegenerative/neuroregenerative processes. Laser microdissection offers a unique opportunity to study molecular regulation in specific immunolabeled cell types. However, an adequate protocol to allow morphological and molecular analysis of rodent spinal cord astrocyte, microglia and motoneurons remains a big challenge. In this paper we present a quick method to immunolabel those cells in flash frozen sections to be used in molecular biology analyses after laser microdissection and pressure catapulting.

MHC class I and II expression in juvenile dermatomyositis skeletal muscle

Objective To assess MHC I and II expressions in muscle fibres of juvenile dermatomyositis (JDM) and compare with the expression in polymyositis (PM), dermatomyositis (DM) and dystrophy. Patients and methods Forty-eight JDM patients and 17 controls (8 PM, 5 DM and 4 dystrophy) were studied. The mean age at disease onset was 7.1 +/- 3.0 years and the mean duration of weakness before biopsy was 9.4 +/- 12.9 months. Routine histochemistry and immunohistochemistry (StreptABComplex/HRP) for MHC I and II (Dakopatts) were performed on serial frozen muscle sections in all patients. Mann-Whitney, Kruskal Wallis, chi-square and Fisher`s exact statistical methods were used. Results MHC I expression was positive in 47 (97.9%) JDM cases. This expression was observed independent of time of disease corticotherapy previous to muscle biopsy and to the grading of inflammation observed in clinical, laboratorial and histological parameters. The expression of MHC I was similar on JDM, PM and DM, and lower in dystrophy. On the other hand, MHC II expression was positive in just 28.2% of JDM cases was correlated to histological features as inflammatory infiltrate, increased connective tissue and VAS for global degree of abnormality (p < 0.05). MCH II expression was similar in DM/PM and lower in JDM and dystrophy, and it was based on the frequency of positive staining rather than to the degree of the MCH II expression. Conclusions MHC I expression in muscle fibres is a premature and late marker of JDM patient independent to corticotherapy, and MHC II expression was lower in JDM than in PM and DM.