Protective effect of the tunneling nanotube-TNFAIP2/M-sec system on podocyte autophagy in diabetic nephropathy.

Podocyte injury leading to albuminuria is a characteristic feature of diabetic nephropathy (DN). Hyperglycemia and advanced glycation end products (AGEs) are major determinants of DN. However, the underlying mechanisms of podocyte injury remain poorly understood. The cytosolic protein TNFAIP2/M-Sec is required for tunneling nanotubes (TNTs) formation, which are membrane channels that transiently connect cells, allowing organelle transfer. Podocytes express TNFAIP2 and form TNTs, but the potential relevance of the TNFAIP2-TNT system in DN is unknown. We studied TNFAIP2 expression in both human and experimental DN and the renal effect of tnfaip2 deletion in streptozotocin-induced DN. Moreover, we explored the role of the TNFAIP2-TNT system in podocytes exposed to diabetes-related insults. TNFAIP2 was overexpressed by podocytes in both human and experimental DN and exposre of podocytes to high glucose and AGEs induced the TNFAIP2-TNT system. In diabetic mice, tnfaip2 deletion exacerbated albuminuria, renal function loss, podocyte injury, and mesangial expansion. Moreover, blockade of the autophagic flux due to lysosomal dysfunction was observed in diabetes-injured podocytes both in vitro and in vivo and exacerbated by tnfaip2 deletion. TNTs allowed autophagosome and lysosome exchange between podocytes, thereby ameliorating AGE-induced lysosomal dysfunction and apoptosis. This protective effect was abolished by tnfaip2 deletion, TNT inhibition, and donor cell lysosome damage. By contrast, Tnfaip2 overexpression enhanced TNT-mediated transfer and prevented AGE-induced autophagy and lysosome dysfunction and apoptosis. In conclusion, TNFAIP2 plays an important protective role in podocytes in the context of DN by allowing TNT-mediated autophagosome and lysosome exchange and may represent a novel druggable target.Abbreviations: AGEs: advanced glycation end products; AKT1: AKT serine/threonine kinase 1; AO: acridine orange; ALs: autolysosomes; APs: autophagosomes; BM: bone marrow; BSA: bovine serum albumin; CTSD: cathepsin D; DIC: differential interference contrast; DN: diabetic nephropathy; FSGS: focal segmental glomerulosclerosis; HG: high glucose; KO: knockout; LAMP1: lysosomal-associated membrane protein 1; LMP: lysosomal membrane permeabilization; MAP1LC3/LC3: microtubule-associated protein 1 light chain 3; PI3K: phosphoinositide 3-kinase; STZ: streptozotocin; TNF: tumor necrosis factor; TNFAIP2: tumor necrosis factor, alpha-induced protein 2; TNTs: tunneling nanotubes; WT: wild type.

African swine fever: A review of cleaning and disinfection procedures in commercial pig holdings.

African swine fever (ASF) is one of the most important diseases in pigs. Since there are no effective vaccines against the virus, farm biosecurity and good farming practices are the only effective tools to prevent the spread of the ASF virus (ASFV) in pig holdings. Hence, an important component of farm biosecurity is the Cleaning and Disinfection (C&D) procedure. Precise indications regarding the ideal disinfectant against ASFV are lacking, but every country has approved and/or authorized a list of biocides effective against ASFV. Lipidic solvents, which destroy the envelope of the virus and commercial disinfectants based on iodine and phenolic compounds are effective in inactivating the ASFV. This review describes the C&D protocol to apply in pig holdings with particular reference to ASFV.

Reversal of albuminuria by combined AM6545 and perindopril therapy in experimental diabetic nephropathy.

BACKGROUND AND PURPOSE: The endocannabinoid (EC) system has been implicated in the pathogenesis of diabetic nephropathy (DN). We investigated the effects of peripheral blockade of the cannabinoid CB1 receptor as an add-on treatment to ACE-inhibition in type 1 diabetic mice (DM) with established albuminuria. EXPERIMENTAL APPROACH: Renal functional parameters (albumin excretion rate, creatinine clearance), tubular injury, renal structure, both EC and CB receptor levels and markers of podocyte dysfunction, fibrosis and inflammation were studied in streptozotocin-induced DM treated for 14 weeks with vehicle, the ACE-inhibitor perindopril (2 mg·kg-1 ·day-1 ), peripherally-restricted CB1 receptor antagonist AM6545 (10 mg·kg-1 ·day-1 ) or both. Treatments began at 8 weeks after diabetes onset, when early DN is established. KEY RESULTS: CB1 receptors were overexpressed in DM and neither perindopril nor AM6545 altered this effect, while both drugs abolished diabetes-induced overexpression of angiotensin AT1 receptors. Single treatment with either AM6545 or perindopril significantly reduced progression of albuminuria, down-regulation of nephrin and podocin, inflammation and expression of markers of fibrosis. However, reversal of albuminuria was only observed in mice administered both treatments. The ability of the combination therapy to completely abolish slit diaphragm protein loss, monocyte infiltration, overexpression of inflammatory markers and favour macrophage polarization towards an M2 phenotype may explain this greater efficacy. In vitro experiments confirmed that CB1 receptor activation directly inhibits retinoic acid-induced nephrin expression in podocytes and IL-4-induced M2 polarization in macrophages. CONCLUSION AND IMPLICATIONS: Peripheral CB1 receptor blockade used as add-on treatment to ACE-inhibition reverses albuminuria, nephrin loss and inflammation in DM.

MicroRNA and Microvascular Complications of Diabetes.

In the last decade, miRNAs have received substantial attention as potential players of diabetes microvascular complications, affecting the kidney, the retina, and the peripheral neurons. Compelling evidence indicates that abnormally expressed miRNAs have pivotal roles in key pathogenic processes of microvascular complications, such as fibrosis, apoptosis, inflammation, and angiogenesis. Moreover, clinical research into innovative both diagnostic and prognostic tools suggests circulating miRNAs as possible novel noninvasive markers of diabetes microvascular complications. In this review, we summarize current knowledge and understanding of the role of miRNAs in the injury to the microvascular bed in diabetes and discuss the potential of miRNAs as clinical biomarkers of diabetes microvascular complications.

Cannabinoid Receptors in Diabetic Kidney Disease.

PURPOSE OF REVIEW: The purpose of this review is to examine and summarize studies assessing the relevance of the endocannabinoid system (ECS) in diabetic kidney disease (DKD). RECENT FINDINGS: Endocannabinoids and endocannabinoid receptors of type 1 (CB1R) and of type 2 (CB2R) are present in the normal kidney. Expression of CB1R and CB2R is altered in experimental DKD. Studies in experimental animals and cultured kidney cells show a beneficial effect of peripheral CB1R blockade and CB2R activation in DKD and an even greater efficacy of a combined treatment. Preclinical studies confirm that both CB1R and CB2R are implicated in the pathogenesis of DKD and may represent novel targets for treatment. However, we need to gain a better understanding of the ECS prior to move to human clinical trial.

Disrupted in schizophrenia 1 (DISC1) L100P mutants have impaired activity-dependent plasticity in vivo and in vitro.

Major neuropsychiatric disorders are genetically complex but share overlapping etiology. Mice mutant for rare, highly penetrant risk variants can be useful in dissecting the molecular mechanisms involved. The gene disrupted in schizophrenia 1 (DISC1) has been associated with increased risk for neuropsychiatric conditions. Mice mutant for Disc1 display morphological, functional and behavioral deficits that are consistent with impairments observed across these disorders. Here we report that Disc1 L100P mutants are less able to reorganize cortical circuitry in response to stimulation in vivo. Molecular analysis reveals that the mutants have a reduced expression of PSD95 and pCREB in visual cortex and fail to adjust expression of such markers in response to altered stimulation. In vitro analysis shows that mutants have impaired functional reorganization of cortical neurons in response to selected forms of neuronal stimulation, but there is no altered basal expression of synaptic markers. These findings suggest that DISC1 has a critical role in the reorganization of cortical plasticity and that this phenotype becomes evident only under challenge, even at early postnatal stages. This result may represent an important etiological mechanism in the emergence of neuropsychiatric disorders.

Neuropsychological effects of the CSMD1 genome-wide associated schizophrenia risk variant rs10503253.

The single-nucleotide polymorphism (SNP) rs10503253, located within the CUB and Sushi multiple domains-1 (CSMD1) gene on 8p23.2, was recently identified as genome-wide significant for schizophrenia (SZ), but is of unknown function. We investigated the neurocognitive effects of this CSMD1 variant in vivo in patients and healthy participants using behavioral and imaging measures of brain structure and function. We compared carriers and non-carriers of the risk 'A' allele on measures of neuropsychological performance typically impaired in SZ (general cognitive ability, episodic and working memory and attentional control) in independent samples of Irish patients (n = 387) and controls (n = 171) and German patients (205) and controls (n = 533). Across these groups, the risk 'A' allele at CSMD1 was associated with deleterious effects across a number of neurocognitive phenotypes. Specifically, the risk allele was associated with poorer performance on neuropsychological measures of general cognitive ability and memory function but not attentional control. These effects, while significant, were subtle, and varied between samples. Consistent with previous evidence suggesting that CSMD1 may be involved in brain mechanisms related to memory and learning, these data appear to reflect the deleterious effects of the identified 'A' risk allele on neurocognitive function, possibly as part of the mechanism by which CSMD1 is associated with SZ risk.

Prospective evaluation of interobserver diagnostic agreement for focal urinary bladder wall abnormalities detected by ultrasound.

INTRODUCTION: The interobserver variability of the sonographic assessment of Focal Bladder Wall Abnormalities (FBWA) between two physicians with different levels of experience was evaluated prospectively. MATERIALS AND METHODS: In the same session the two operators examined independently the urinary bladder of 87 consecutive patients (Mean Age 68 yrs; range 33-80; 75 Males; 15 Females) who underwent cystoscopy within 1-2 days. Before beginning the study, the two observers standardized the process for US exam. Weighted kappa statistics were used to determine the degree of agreement as to the presence, size, location and number of FBWA detected at cystoscopy. RESULTS: the highest level of agreement (k = 1) was reached on the presence of FBWA, whereas a substantial agreement was reached on size (k = 0.78), number (k = 0.72) and location (k = 0.62) of FBWA. CONCLUSIONS: In this study the gap in US experience between two operators did not emerge in assessing FBWA, provided a standardized technical approach is pursued.

Swine vesicular disease in northern Italy: diffusion through densely populated pig areas.

At the end of 2006, a recrudescence of swine vesicular disease (SVD) was recorded in Italy and the disease spread widely throughout the northern regions. Lombardy, a densely populated pig area, was most affected and the presence of the disease caused heavy economic losses to the entire pig industry. Although SVD is considered only moderately contagious, the epidemic in the north was characterised by a rapid spread of the condition. Numerous difficulties were encountered in eradicating it. Over the past decade, there has been a significant increase in the population of pigs in Lombardy, concentrated mainly in a few areas which were the most severely affected during the 2006 to 2007 SVD epidemic. Increases in both the pig population and animal movements, combined with weak biosecurity measures, increased the spread rate of the disease and hampered eradication activities.

Swine vesicular disease surveillance and eradication activities in Italy.

Swine vesicular disease (SVD) was first observed in Italy in 1966, and was initially diagnosed as foot and mouth disease (FMD). The causative agent of SVD was classified as an Enterovirus within the family Picornaviridae. It was included in the list of diseases notifiable to the World Organisation for Animal Health (OIE) because of the similarity of its lesions to those produced by FMD; however SVD is often mild in nature and may infect pigs subclinically. During the last decade SVD has been persistently reported in Italy, and surveillance and eradication activities are in place. The central and northern parts of Italy have been designated SVD free since 1997, while the southern regions have not achieved disease-free status. However, occasional outbreaks of SVD have occurred in central and northern Italy and have been eradicated using rigorous control measures. Most recent SVD outbreaks in Italy have been subclinical; SVD can rarely be diagnosed now on the basis of clinical signs and it is necessary to use laboratory diagnosis. This paper examines the epidemiology of SVD in Italy, and considers the measures adopted in Europe for SVD control on the basis of current knowledge of the disease.

Tension-free transobturator approach for female stress urinary incontinence.

AIM: UraTape is a new sling for female stress urinary incontinence (SUI) inserted via a transobturator percutaneous approach. We report the safety, feasibility and short-term results of this new surgical procedure. METHODS: UraTape (Porgès-Mentor) is a non-elastic polypropylene tape, with a silicone coated central part. The sling is placed tension-free under the mid-urethra and extended through the obturator foramen bypassing the posterior face of the ischiopubic ramus. From September 2002 to May 2003, 80 females affected by SUI associated with urethral hypermobility and without severe uro-genital prolapse (with cystocele 1st grade) received UraTape. Mean age was 56 (39-79) years. Sixty-two out of 80 had a positive Q tip test; 16/80 had recurrent incontinence after Burch or colpoplasty according to Kelly; 22/60 had mixed incontinence. Preoperative evaluations included: complete history and physical examination, urinalysis, urodynamic investigations, abdominal and pelvic ultrasound. Quality of life assessment was carried out pre- and postoperatively. RESULTS: Mean operative time was 16 (11-36) minutes. No major intraoperative complications were observed. One bladder neck laceration occurred and was treated intraoperatively. No cystoscopy was performed. Mean hospital postoperative stay time was 1.1 (1-6) days. All patients were examined periodically at 7, 30 and 90 days from intervention (mean follow-up 4 months, 1-8). There was no urethral erosion. One vaginal erosion with inguinal abscess was diagnosed and treated without removing the sling. Two de novo urge incontinence was observed. The objective and subjective cure rates were 92% and 97%; 96% expressed good quality of life (satisfied/very satisfied). CONCLUSION: This procedure is a safe, effective new technique for the treatment of female SUI. The easy technique, the short learning curve and the very high grade of satisfaction of the patients show that this approach is based upon effective anatomical and physiological criteria. Further follow-up is necessary in order to evaluate urodynamic changes and overall satisfaction.

Contextual Assessment of the Maternity Experience: development of an instrument for cross-cultural research.

BACKGROUND: There is evidence that stressors may trigger the onset of a depressive episode in vulnerable women. A new UK interview measure, the Contextual Assessment of the Maternity Experience (CAME), was designed to assess major risk factors for emotional disturbances, especially depression, during pregnancy and post-partum. AIMS: With in the context of a cross-cultural study, to establish the usefulness of the CAME, and to test expected associations of the measure with characteristics of the social context and with major or minor depression. METHOD: The CAME was administered antenatally and postnatally in ten study sites, respectively to 296 and 249 women. Affective disorder throughout pregnancy and up to 6 months postnatally was assessed by means of the Structured Clinical Interview for DSM-IVAxis I Disorders. RESULTS: Adversity, poor relationship with either a partner or a confidant, and negative feelings about the pregnancy all predicted onset of depression during the perinatal period. CONCLUSIONS: The CAME was able to assess major domains relevant to the psychosocial context of the maternity experience in different cultures. Overall, the instrument showed acceptable psychometric properties in its first use in different cultural settings.

A locus for migraine without aura maps on chromosome 14q21.2-q22.3.

Migraine is a common and disabling neurological disease of unknown origin characterized by a remarkable clinical variability. It shows strong familial aggregation, suggesting that genetic factors are involved in its pathogenesis. Different approaches have been used to elucidate this hereditary component, but a unique transmission model and causative gene(s) have not yet been identified. We report clinical and molecular data from a large Italian pedigree in which migraine without aura (MO) segregates as an autosomal dominant trait. After exclusion of any association between MO and the known familial hemiplegic migraine and migraine with aura loci, we performed a genomewide linkage analysis using 482 polymorphic microsatellite markers. We obtained significant evidence of linkage between the MO phenotype and the marker D14S978 on 14q22.1 (maximum two-point LOD score of 3.70, at a recombination fraction of 0.01). Multipoint parametric analysis (maximum LOD score of 5.25 between markers D14S976 and D14S978) and haplotype construction showed strong evidence of linkage in a region of 10 cM flanked by markers D14S1027 and D14S980 on chromosome 14q21.2-q22.3. These results indicate the first evidence of a genetic locus associated with MO on chromosome 14.

Genetic mapping of a susceptibility locus for disc herniation and spastic paraplegia on 6q23.3-q24.1.

It has been suggested that a genetic factor(s) or a familial predisposition may contribute to the clinical manifestations of disc herniation; moreover, no genetic linkage between spinal disc herniation and spastic paraplegia has ever been described. A family with consanguineous parents and four of eight sibs affected by multiple disc herniations and spastic paraplegia was clinically and genetically analysed. Surgery caused partial improvement in all of them. After the exclusion of type II collagen and vitamin D receptor genes and the recessive loci for HSPs, a genome wide search was performed with about 500 fluorescent markers. Positive lod score values were obtained for chromosome 6q22.31-q24.1, with evidence of three homozygous intervals. The maximum multipoint lod score of 3.28 was obtained in only one interval, between markers D6S1699 and D6S314. On the whole, a susceptibility locus for disc herniation and autosomal recessive spastic paraplegia was found on chromosome 6q23.3-q24.1. This is the first time that disc herniation and the associated neurological syndrome has been linked to a human chromosomal region.

Spatial risk factors related to outbreaks of contagious bovine pleuropneumonia in northern Italy (1990-1993).

In October 1990, an outbreak of contagious bovine pleuropneumonia (CBPP) was reported in Italy after an absence of approximately one century. Since October 1990, ninety-four outbreaks have occurred in Italy, of which forty-seven were concentrated in three areas of northern Italy (Lombardy region). The disease was eradicated in September 1993. The data used for the analysis were obtained from the epidemiological investigations undertaken during the outbreaks of 1990-1993. The unit of interest for the analysis is the farm. Spatial segregation of infected and uninfected farms within the study area was determined through the Pielou index of segregation. Data from herds within the same set of outbreaks were analysed through logistic regression to identify factors which could be used to discriminate between infected and uninfected herds. The study indicated a clear spatial segregation between infected and uninfected herds. The results of the analysis do not indicate the mode of disease spread. However, the study demonstrated that both aerosol and indirect transmission of the infection could have occurred, as previously documented in Africa. The possibility that indirect transmission played a prominent role in the spread of CBPP in the region of Lombardy is a completely new suggestion. Further studies are required to understand the epidemiology of CBPP in regions with intensive farming and a relatively cold climate. In particular, three aspects require consideration; firstly, animal movement among neighbouring herds could produce a pattern of disease similar to the one produced by indirect transmission (this possibility was excluded in the epidemiological outbreak investigations conducted by field veterinarians). Secondly, the methods of spatial analysis used in this study have not been previously used in the field of veterinary epidemiology. Further validation of the efficacy of these methods is thus required. Thirdly, the epidemiology of CBPP under conditions of high animal density and a relatively cool climate, as encountered in Lombardy, requires further investigation.

Implementation of a system for the regional management of animal health emergencies.

A telematic system to support decisions and operations in case of animal health emergencies has been designed and implemented in the Abruzzo region of Italy. The system aims to improve decision-making by Veterinary Services in the event of an outbreak of exotic disease. The system has been tested, first by a simulated outbreak of foot and mouth disease, and then during an outbreak of swine vesicular disease. Critical problems were detected and corrected in both cases.

Management of animal health emergencies: general principles and legal and international obligations.

The Eighth Round of the General Agreement on Tariffs and Trade led to the creation of the World Trade Organization and to the adoption of the Sanitary and Phytosanitary (SPS) Agreement, thus considerably changing the rules of international trade in animals and animal products. Animal health measures may result in trade restrictions, but governments accept that these restrictions may sometimes be necessary and appropriate to ensure food safety and animal health protection. The SPS Agreement acknowledges the rights of governments to adopt measure to protect human, animal and plant health. To ensure effective animal health protection, without unjustifiable discrimination, the operational procedures of Veterinary Services must be standardised, especially those concerning disease notification, epidemiological information, certification for international trade and management of animal health emergencies. Veterinary Services must be further supported by a proper legislative framework and adequate financial resources.

Sensitivity and specificity of serological and bacteriological tests for contagious bovine pleuropneumonia.

In 1990 an outbreak of contagious bovine pleuropneumonia (CBPP) occurred in Italy. Subsequent surveillance for CBPP was based on random sampling in bovine herds, serological controls on all animals moved from the herd of origin and controls on slaughtered animals. Official tests employed were the complement fixation test (CFT) and bacteriological isolation and typing. A total of 33,856 serum samples collected from herds in CBPP-free regions were used to define CFT specificity, while samples from 595 animals from infected herds were employed to define the sensitivity. Ninety-nine animals from three infected herds were used to estimate the sensitivity of the isolation technique. Results showed the specificity of CFT (threshold +1:10) to be 98% and sensitivity to be 63.79%. The sensitivity of the test did not change significantly, regardless of whether the lesions were caused by acute or chronic infection. The sensitivity of the isolation technique was 54.1%.

Simultaneous chiral separation of 3,4-methylenedioxymethamphet- amine, 3-4-methylenedioxyamphetamine, 3,4-methylenedioxyethylam- phetamine, ephedrine, amphetamine and methamphetamine by capillary electrophoresis in uncoated and coated capillaries with native beta-cyclodextrin as the chiral selector: preliminary application to the analysis of urine and hair.

The importance of the chiral analysis of amphetamine-related substances in both clandestine preparations and biological samples is widely recognized. For this purpose, capillary electrophoresis was successfully applied by several authors, but only few reports concerned ring-substituted amphetamines, which represent the main components of "ecstasy", a widely abused "recreational" substance. In the present work, the simultaneous chiral analysis of ephedrine, amphetamine, methamphetamine, 3,4-methylenedioxymethamphetamine (MDMA), 3-4-methylenedioxyamphetamine (MDA) and 3,4-methalenedioxyethylamphetamine (MDE) is reported, by using capillary electrophoresis with native beta-cyclodextrin (15 mM) as the chiral selector. After preliminary tests at different pH values (phosphate buffer 100 mM, pH 2.5-9.0) and with bare or coated fused-silica capillaries, the optimized conditions were: pH 2.5 phosphate, uncoated capillary (45 cm x 50 microm inner diameter), potential 10 kV. Detection was either by fixed wavelength (200 nm) or multiwavelength (190-400 nm) UV absorbance. Under these conditions, good resolution was obtained for all the analytes, with excellent chiral selectivity and efficiency. The sensitivity for the individual enantiomers was better than 0.2 microg/mL, analytical precision was characterized by relative standard deviation values < 0.8% (< or = 0.15% with internal standardization) for migration times intra-day and < 2.0% (< or = 0.54% with internal standardization) day-to-day; linearity, in the range 0.156-40 microg/mL, and accuracy were also satisfactory. After a simple liquid-liquid extraction, urine samples could be analyzed with a sensitivity well below the recommended NIDA cut-off of 500 ng/mL. For hair samples, it was necessary to increase the sensitivity by applying a field-amplified sample stacking procedure, which allowed the chiral determination of MDA, MDMA and MDE at concentrations occurring in real samples from ecstasy users, with the possibility of recording UV spectra of the peaks.