Cdk12 maintains the integrity of adult axons by suppressing actin remodeling.

The role of cyclin-dependent kinases (CDKs) that are ubiquitously expressed in the adult nervous system remains unclear. Cdk12 is enriched in terminally differentiated neurons where its conical role in the cell cycle progression is redundant. We find that in adult neurons Cdk12 acts a negative regulator of actin formation, mitochondrial dynamics and neuronal physiology. Cdk12 maintains the size of the axon at sites proximal to the cell body through the transcription of homeostatic enzymes in the 1-carbon by folate pathway which utilize the amino acid homocysteine. Loss of Cdk12 leads to elevated homocysteine and in turn leads to uncontrolled F-actin formation and axonal swelling. Actin remodeling further induces Drp1-dependent fission of mitochondria and the breakdown of axon-soma filtration barrier allowing soma restricted cargos to enter the axon. We demonstrate that Cdk12 is also an essential gene for long-term neuronal survival and loss of this gene causes age-dependent neurodegeneration. Hyperhomocysteinemia, actin changes, and mitochondrial fragmentation are associated with several neurodegenerative conditions such as Alzheimer's disease and we provide a candidate molecular pathway to link together such pathological events.

Anal endosonography and fistulography for fistula-in-ano.

PURPOSE: In this study, a comparison was made of the accuracy and clinical usefulness of anal endosonography and fistulography in the preoperative classification of fistulas-in-ano. MATERIALS AND METHODS: A total of 113 patients with a clinical diagnosis of cryptoglandular fistula-in-ano who were awaiting surgery were included in this retrospective review. Patients were preoperatively investigated by anal endosonography and/or modified fistulography by inserting a Foley catheter into the rectum and a metal ring close to the anus. The catheter and ring served as radiopaque anal markers. Fistula classification obtained by the two diagnostic modalities was compared with surgical classification as the criterion standard. RESULTS: Endoanal ultrasound and fistulography identified 82.8% and 100% of primary tracks, 79% and 74.2% of internal openings, 98% and 91.8% of secondary tracks and 92.9% and 87.8% of abscesses, respectively. CONCLUSIONS: Anal endosonography and fistulography with radiopaque markers are important complements to surgical exploration for investigating anal sepsis and may be of value to the surgeon in planning a therapeutic strategy.

Accelerated diabetic glomerulopathy in galectin-3/AGE receptor 3 knockout mice.

Several molecules were shown to bind advanced glycation end products (AGEs) in vitro, but it is not known whether they all serve as AGE receptors and which functional role they play in vivo. We investigated the role of galectin-3, a multifunctional lectin with (anti)adhesive and growth-regulating properties, as an AGE receptor and its contribution to the development of diabetic glomerular disease, using a knockout mouse model. Galectin-3 knockout mice obtained by gene ablation and the corresponding wild-type mice were rendered diabetic with streptozotocin and killed 4 months later, together with age-matched nondiabetic controls. Despite a comparable degree of metabolic derangement, galectin-3-deficient mice developed accelerated glomerulopathy vs. the wild-type animals, as evidenced by the more pronounced increase in proteinuria, extracellular matrix gene expression, and mesangial expansion. This was associated with a more marked renal/glomerular AGE accumulation, indicating it was attributable to the lack of galectin-3 AGE receptor function. The galectin-3-deficient genotype was associated with reduced expression of receptors implicated in AGE removal (macrophage scavenger receptor A and AGE-R1) and increased expression of those mediating cell activation (RAGE and AGE-R2). These results show that the galectin-3-regulated AGE receptor pathway is operating in vivo and protects toward AGE-induced tissue injury in contrast to that through RAGE.

Increased retinal endothelial cell monolayer permeability induced by the diabetic milieu: role of advanced non-enzymatic glycation and polyol pathway activation.

BACKGROUND: Increased vascular permeability could be involved in the pathogenesis of diabetic retinopathy. The present study was aimed at assessing whether high glucose concentrations can impair retinal endothelial cell barrier function directly, irrespective of changes in other determinants of permeability, and the role of non-enzymatic glycation and polyol pathway activation in these alterations. METHODS: Bovine retinal endothelial cells (BREC) were exposed for various periods to high glucose vs iso-osmolar mannitol and normal glucose containing media+/-agents mimicking or inhibiting advanced glycation end product (AGE) formation and polyol pathway activation. Monolayer permeability was assessed by measuring the transendothelial passage of (125)I-labeled proteins. RESULTS: Permeability increased significantly (up to +70%) in BREC exposed to high glucose, but not to mannitol, for 1-30 days, vs normal glucose control cells. Exposure to AGE-modified bovine serum albumin (BSA) (> or = 90%) and, to a lesser extent, sorbitol (+28%) mimicked the high glucose effect. The AGE formation and nitric oxide synthase (NOS) inhibitor aminoguanidine significantly reduced (by 60%) changes induced by 30-day exposure to high glucose, whereas methylguanidine, which inhibits only NOS activity, did not affect permeability. Aldose reductase or sorbitol dehydrogenase inhibitors decreased (by approximately 40%) the enhanced leakage produced by 1-day, but not 30-day, incubation in high glucose. CONCLUSIONS: The present results indicate that high glucose is capable of impairing retinal endothelial cell barrier function directly and that non-enzymatic glycation and polyol pathway activation may mediate these changes, with AGEs participating in the long-term alterations and increased flux through the sorbitol pathway in the short-term effect.

Role of galectin-3 as a receptor for advanced glycosylation end products.

The advanced glycosylation end product (AGE)-binding proteins identified so far include the components of the AGE-receptor complex p60, p90 and galectin-3, receptor for advanced glycosylation end products (RAGE), and the macrophage scavenger receptor types I and II. Galectin-3 interacts with beta-galactoside residues of several cell surface and matrix glycoproteins through the carbohydrate recognition domain and is also capable of peptide-peptide associations mediated by its N-terminus domain. These structural properties enable galectin-3 to exert multiple functions, including the modulation of cell adhesion, the control of cell cycle, and the mRNA splicing activity. Moreover, in macrophages, astrocytes, and endothelial cells, galectin-3 has been shown to exhibit a high-affinity binding for AGEs; the lack of a transmembrane anchor sequence or signal peptide suggests that it associates with other AGE-receptor components rather than playing an independent role as AGE-receptor. In tissues that are targets of diabetic vascular complications, such as the mesangium and the endothelium, galectin-3 is not expressed or only weakly expressed under basal conditions, at variance with p90 and p60 but becomes detectable with aging and is induced or up-regulated by the diabetic milieu, which only slightly affects the expression of p90 or p60. This (over)expression of galectin-3 may in turn modulate AGE-receptor-mediated events by modifying the function of the AGE-receptor complex, which could play a role in the pathogenesis of target tissue injury. Up-regulated galectin-3 expression may also exert direct effects on tissue remodeling, independently of AGE ligands, by virtue of its adhesive and growth regulating properties.

Do men and women follow different trajectories to reach extreme longevity? Italian Multicenter Study on Centenarians (IMUSCE).

Gender accounts for important differences in the incidence and prevalence of a variety of age-related diseases. Considering people of far advanced age, demographic data document a clear-cut prevalence of females compared to males, suggesting that sex-specific mortality rates follow different trajectories during aging. In the present investigation, we report data from a nationwide study on Italian centenarians (a total of 1162 subjects), and from two studies on centenarians living in two distinct zones of Italy, i.e., the island of Sardinia (a total of 222 subjects) and the Mantova province (Northern Italy) (a total of 43 subjects). The female/male ratio was about 2:1 in Sardinia, 4:1 in the whole of Italy, and about 7:1 in the Mantova province. Thus, a complex interaction of environmental, historical and genetic factors, differently characterizing the various parts of Italy, likely plays an important role in determining the gender-specific probability of achieving longevity. Gender differences in the health status of centenarians are also reported, and an innovative score method to classify long-lived people in different health categories, according to clinical and functional parameters, is proposed. Our data indicate that not only is this selected group of people, as a whole, highly heterogeneous, but also that a marked gender difference exists, since male centenarians are less heterogeneous and more healthy than female centenarians. Immunological factors regarding the age-related increase in pro-inflammatory status, and the frequency of HLA ancestral haplotypes also show gender differences that likely contribute to the different strategies that men and women seem to follow to achieve longevity. Concerning the different impact of genetic factors on the probability of reaching the extreme limits of the human life-span, emerging evidence (regarding mtDNA haplogroups, Thyrosine Hydroxilase, and IL-6 genes) suggests that female longevity is less dependent on genetics than male longevity, and that female centenarians likely exploited a healthier life-style and more favorable environmental conditions, owing to gender-specific cultural and anthropological characteristics of the Italian society in the last 100 years.

The diabetic milieu modulates the advanced glycation end product-receptor complex in the mesangium by inducing or upregulating galectin-3 expression.

Nonenzymatic glycation has been implicated in the pathogenesis of the dysregulated tissue remodeling that characterizes diabetic glomerulopathy, via the formation of advanced glycation end products (AGEs) and their binding to cell surface receptors. Several AGE-binding proteins have been identified so far, including p60, p90, and the adhesive and growth-regulating lectin galectin-3 (Gal-3), the components of the so-called AGE-receptor complex. This study aimed to evaluate the mesangial expression of the AGE-receptor complex and its modulation by the diabetic milieu, both in vivo, in non-diabetic versus streptozotocin-induced diabetic rats, and in vitro, in mesangial cells exposed to either normal glucose (NG) levels (5.5 mmol/l), as compared with high glucose (HG) levels (30 mmol/l) and iso-osmolar mannitol (M), or to native bovine serum albumin (BSA), as compared with glycated BSA with AGE formation (BSA-AGE) and glycated BSA in which AGE formation was prevented by aminoguanidine (BSA-AM). In vivo, Gal-3 protein and mRNA were not detectable in glomeruli from nondiabetic rats until 12 months after initiating the study. On the contrary, in diabetic rats, Gal-3 expression was observed at 2 months of disease duration, and it increased thereafter. Both p60 and p90 immunoreactivities were observed at the glomerular level with slightly increased expression of p90, but not p60, in diabetic versus nondiabetic animals. In vitro, Gal-3 was not detectable in mesangial cells cultured in NG (although it became evident after a certain number of passages in culture), whereas Gal-3 was detectable in cells grown on BSA. Prolonged exposure (2-4 weeks) of mesangial cells to HG but not to M, as well as growing cells on BSA-AGE and, to a lesser extent, BSA-AM, induced or significantly increased the expression of Gal-3, both protein (up to 2.65-fold) and mRNA (up to 3.10-fold) and its secretion in the medium (by approximately 50%). Both p60 and p90 were demonstrated in mesangial cells under NG conditions, and the expression of p90, but not p60, was upregulated by approximately 20% by HG or BSA-AGE. These results indicate that 1) under basal conditions, Gal-3, unlike p90 and p60, is not detectable in the mesangium but becomes expressed with aging and 2) the diabetic milieu induces or upregulates Gal-3 production, whereas it increases only slightly the expression of p90, but not p60. Gal-3 expression or overexpression may modulate the AGE-receptor-mediated events by modifying the function of the AGE-receptor complex. Additionally, it may exert direct effects on tissue remodeling by virtue of its adhesive and growth-regulating properties.

Dynamic imaging of the normal pelvic floor.

PURPOSE: To provide quantitative data by a modern cross-sectional imaging technique (CT) for defining normal physiological values of pelvic floor structures. PATIENTS AND METHODS: Twenty seven subjects, 7 males, 20 females, aged 20-75 yrs (mean 46.3 +/- 5 yrs) without pelvic floor or defection dysfunction underwent Direct Coronal (DC) CT scanning of the pelvis with the patient seated instead of lying. Scans obtained at rest and on straining were compared by bony landmarks. Three anatomical compartments, i.e. anterior, middle and posterior, were identified by two planes drawn tangential to the ischial foramina and the ischial tuberosities, respectively. Measurements of (1) Levator ani muscle length (mm); (2) Levator-anal angle (degrees); (3) Rectal floor-to-ischial line distance (mm) and (4) Supra/Infralevator spaces (square cm) were independently performed twice by two radiologists. The statistical analysis included calculation of intra and interobserver agreement (correlation coefficient). The differences between the means of the resting and straining values from each compartment (Student's t test) and the correlation between parameters (Pearson's coefficient) to evaluate whether resting values allowed a prediction of those on straining were determined. RESULTS: DC scans of diagnostic quality were obtained in all but two patients (92.5%). Both intra- and interobserver agreement indices were always greater than 80% (except for a 0.63 value by one observer obtained in the infralevator space from the anterior compartment at rest). A significant difference between the resting and straining values of all parameters was noted in the three compartments. At rest the levator ani muscle length was significantly shorter and the supralevator space smaller in the posterior compartment (48.3 +/- 7.9 mm vs 48.8 +/- 7 mm vs 42.6 +/- 9.4 mm, P < 0.05 and 70.6 +/- cm2 vs 66.9 +/- 11.5 cm2 vs 27.2 +/- 4.8 cm2, P < 0.01 anterior, middle and posterior respectively). On straining, these two parameters increased by +42% and +17.8%, respectively, in the same compartment, while the most pronounced variation of the infralevator space occurred in the middle compartment (-51.1%). The increase in the supralevator space correlated with a decrease in the rectal floor-to-ischial line distance and widening of the levator-anal angle (r = -0.64, P < 0.01 and 0.48, P < 0.05, respectively). A close correlation between resting and straining values was observed in all parameters, especially in the supralevator space in the three compartments (r = 0.82, 0.93 and 0.88, P < 0.01). CONCLUSIONS: Direct Coronal CT scanning showed that on straining the posterior component of the levator ani muscle, i.e. the coccygeus muscle, undergoes "physiological overstretching" and the supralevator space acts as a "compliant cavity", whose behaviour can be predicted at rest.

An EEG power index (eyes open vs. eyes closed) to differentiate Alzheimer's from vascular dementia and healthy ageing.

EEG differential power patterns between Alzheimer's (AD, 50 patients) and vascular (VaD, 37 patients) dementia and between these two and 36 healthy ageing subjects, were studied in the 6.5-12 Hz band of the ongoing EEG recorded during the rest eyes closed (REC) and eyes open (REO) conditions. From the EEGs (16 electrodes, 10-20 international system except for Fz, Cz, Pz), a 6.5-12 Hz band, wider than the alpha range (alpha-like), was chosen and processed to include the highest theta frequencies characterising the occipital dominant activity in dementia. A global and occipital EEG Power Index (PI) was calculated and used considering the absolute powers during REC and REO. The MANOVA was used to compare the figures. Bearing in mind that the higher the PI value the greater the difference between the 6.5-12 Hz EEG band powers of REC vs. REO, the results were as follows: (i) in the patients with Alzheimer's and vascular dementia the global and occipital PIs were significantly lower than those in controls; (ii) in the patients with Alzheimer's dementia the same PIs were significantly lower that those of the patients with VaD; (iii) healthy elderly subjects showed significantly lower powers in the 6.5-12 Hz frequencies at T5 and O1 in REO as compared to dementia patients. The pathophysiological implications and the clinical applications of these results are discussed.

[The 3rd national workshop on defecography: the functional radiology of (neo) rectal ampullae (ileal reservoir, colo-anal anastomosis, continent perineal colostomy)]. / III workshop nazionale sulla defecografia: la fisioradiologia delle (neo)ampolle rettali (reservoir ileale, colo-ano-anastomosi, colostomia perineale continente).

A survey was made in 13 Italian centers with a questionnaire concerning the (a) indications, (b) postoperative complications, (c) functional results and (d) diagnostic imaging modalities related to the making of an ileal or colonic (neo) rectum. Ulcerative colitis (100%), familial polyposis (61.5%) and Crohn's disease (15.3%) were the most common indications for an ileal pouch; rectal cancer (7.96%), chronic inflammatory diseases (15.3%), diverticulosis, rectal prolapse, redundant colon and imperforate anus (7.6% each) were the most common indications for a colonic pouch. Postoperative complications included pelvic abscess (14%), sinus tract/dehiscence (10%) and bowel obstruction (9%). When compared with the S and W variants, the J-shaped ileoanal pouch proved superior because urgency and fecal retention rates were lower (18.4% vs. 44.4% and 23% vs. 28.6%, p < 0.01 and p < 0.05, respectively), despite slightly more frequent staining episodes (15.8% vs. 11.1%; p < 0.05). As for colonic ampullae, fecal retention and provoked evacuation were more frequent in the J pouch and after gracileplasty; urgency and incontinence in the straight colo-anal anastomosis (33.3% vs. 22.2% and 41.6% vs. 33.3%, respectively). The functional outcome was assessed by anal endosonography (available in 4/13 centers), defecography and anorectal manometry. Abnormal findings included: (a) reduced capacity, barium leakage, anal gaping, sphincter damage (urgency and incontinence); (b) barium retention, pouch dilatation, split evacuation, knobs and strictures (fecal retention).

Effect of lonidamine on the mitochondrial potential in situ in Ehrlich ascites tumor cells.

The effect of lonidamine (LND) on the mitochondrial membrane potential, in situ, of Ehrlich ascites tumour cells was investigated by using the safranine method. LND, because of its ability to inhibit electron transport from endogenous substrates to respiratory carriers, induced a de-energization of mitochondria. Addition of glucose to rotenone-treated cells induced mitochondrial membrane potential as shown by the spectral shift similar to that which occurred upon energization of mitochondria. The build-up of membrane potential in rotenone-treated cells was due to glycolytically-generated ATP because the response to glucose was abolished by LND which inhibited the glycolysis of neoplastic cells by affecting the mitochondria bound hexokinase.

The retinoblastoma gene product in acute myeloid leukemia: a possible involvement in promyelocytic leukemia.

The retinoblastoma susceptibility gene in leukemia and lymphoma has been investigated using different approaches involving either gene or protein analysis. In this study, a novel method, which evaluates the functional status of the retinoblastoma gene product by a binding assay to an in vitro-translated viral oncoprotein, has been applied to leukemic cells from acute myeloid leukemia patients. One hundred twenty-two cases were considered, and 42 of them were also analyzed by Western blot. Results obtained with the two methods were comparable, with the exception of few cases, where the retinoblastoma protein appeared detectable but unable to bind to the viral oncoprotein. The retinoblastoma protein has been found defective mostly in the M3 promyelocytic subtype.

Lipoprotein (a) and vascular disease in diabetic patients.

In order to assess the potential role of lipoprotein (a) as a risk factor for cardiovascular disease in diabetes mellitus, plasma concentrations were measured in a large group (n = 500) of non-insulin-dependent (NIDDM, n = 355) and insulin-dependent (IDDM, n = 145) patients. Concentrations of lipoprotein (a) were compared in diabetic patients with (n = 153) or without (347) documented vascular disease (ischaemic heart disease, peripheral vascular disease or macroangiopathy). They were significantly higher (p < 0.05) in patients with ischaemic heart disease (mean [interquartile range] 15.5 (5.0-38.0) vs 9.0 (4.5-26.0) mg/dl) or macroangiopathy (13.0 (5.0-38.0) vs 9.0 (4.0-25.0) mg/dl) compared to patients without manifestations of vascular disease. In addition, stepwise logistic regression analysis identified lipoprotein (a) levels > or = 30 mg/dl as being independently associated with the presence of cardiovascular disease. Lipoprotein (a) was an independent risk factor for ischaemic heart disease and macroangiopathy in this group of IDDM and NIDDM patients.

Blood pressure self-measurement in the elderly: differences between automatic and semi-automatic systems.

Arterial hypertension is the most common cardiovascular risk factor in the elderly. Its clinical control emphasises the problem of the systems used for monitoring: clinical measurement by the physician, home self-monitoring, ambulatory monitoring, etc. In particular, in the elderly population, the self-monitoring of blood pressure can present further problems associated with their situation. In our study we evaluated, in an elderly population, the differences in the self-recording of blood pressure with automatic and semi-automatic equipment using a mercury sphygmomanometer by a physician as a 'gold standard' control. We studied 28 elderly subjects using a rigid protocol for the self-measurement of their blood pressure. Our results show that automatic equipment is significantly more precise and easier to use than semiautomatic equipment in home self-measurement of blood pressure in elderly people.

Apolipoprotein E polymorphism as a risk factor for vascular disease in diabetic patients.

OBJECTIVE: To examine the prevalence of cardiovascular disease in diabetic patients as a function of apolipoprotein (apo) E polymorphism. RESEARCH DESIGN AND METHODS: The apo E phenotypes and plasma lipid, lipoprotein, and apo levels were determined for 517 Italian diabetic patients. The prevalence of cardiovascular disease (defined as ischemic heart disease [HD] and/or peripheral vascular disease and/or cerebrovascular disease) was assessed as a function of apo E polymorphism at entry and after 4 years. RESULTS: The occurrence of vascular disease did not differ significantly between diabetic patients in the various categories of apo E phenotype either at entry into the study or after 4 years. When expressed as a percentage of patients with disease, we observed--for E2, E3, and E4 carriers, respectively--at entry: IHD, 20.0% (n = 14), 21.0% (n = 79), and 21.5% (n = 14); and macroangiopathy, 24.3% (n = 17), 29.3% (n = 110), and 24.6% (n = 16). Apo E polymorphism did not make a significant contribution to multiple logistic regression models designed to identify the factors associated with the occurrence of vascular disease in diabetic patients. CONCLUSION: Apo E polymorphism and, notably, the apo E4 allele cannot be universally considered as a particular risk factor for cardiovascular disease in diabetic patients.

The influence of age and sex on nutritional parameters in subjects aged 60 years and over.

The nutritional assessment of the elderly shows several interpretative difficulties due to the lack of standard parameters. Moreover chronic age-related diseases can interfere with the physiological nutritional status. Anthropometric (triceps skinfold, arm muscle area, total body muscle mass, fat mass and Body Mass Index (BMI)), biochemical (serum prealbumin, transferrin, ceruloplasmin, total protein and albumin) and immunological (serum lymphocytes) parameters were measured in 583 out-patients aged 60 years or over selected on the basis of clinical and biochemical criteria and with BMI /= 75) for each sex. The F-test analysis for all anthropometric parameters except BMI showed significant differences with respect to age (P < 0.05) and sex (P < 0.05). Among biochemical parameters, prealbumin showed a significant difference for age (P < 0.05) and sex (P < 0.05) (males, 30.3 +/- 8.2; females, 29.1 +/- 7.5) while ceruloplasmin showed a significant difference for sex only (P < 0.05) (males, 40.9 +/- 9.3; females, 43.8 +/- 8.2). When the biochemical mean values obtained in this study were compared with those utilized in the daily routine of the hospital central laboratory, ceruloplasmin and prealbumin resulted in significantly higher (P < 0.05) while total protein and albumin were significantly lower values (P < 0.05).

Permanent cardiac pacing and thromboembolic risk in elderly patients.

In patients undergoing permanent cardiac pacing, the maintenance of atrial contractility is important to ensure adequate ventricular filling and to guarantee an optimal ventricular ejection capacity. The appropriate pacing mode, assuring a suitable mechanical atrioventricular coupling, prevents the onset of atrial fibrillation and contributes to reduction of the risk of subsequent systemic and pulmonary thromboembolic episodes. We examined 461 patients (266 males and 195 females, aged between 52 and 97 years, average age 76.5 +/- 18) paced for conduction disturbances of various degrees and etiology. Of them, 323 patients received ventricular demand pacemaker (VVI group, average age 77.9 years); 138 underwent dual chamber pacing (DCP group, average age 75.2 years), 117 of the latter received universal demand pacing (DDD) and 21 atrial synchronous ventricular demand pacing (VDD). The patients were subsequently divided into two age-groups: Group A (/= 75 years, 287 patients). According to pacing mode and successive development of stable atrial fibrillation (AF), we analysed the occurrence of systemic and/or pulmonary thromboembolic episodes and the incidence of fatal events. During our study, performed from January 1986 to August 1993, 70 embolic episodes were observed in the VVI group and six in the DCP group. Eighty-four patients with VVI units developed AF during follow-up, compared with only five patients in the DCP group. Our data indicate that VVI patients have a higher incidence of AF, embolic complications and cerebrovascular mortality, in comparison with the DCP group. VVI pacing should be avoided, especially in older patients, when atrial rhythmical activity is present.