Diagnosis of leprosy in a Nigerian migrant: implementation of surveillance measures in the current migration context.

BACKGROUND: In Italy, leprosy diagnosis is reported in immigrants from endemic countries or Italians who have stayed in endemic areas. We report the first leprosy case to be observed in a migrant from Nigeria in the Rimini district (Emilia-Romagna, Northern Italy). METHODS: After describing the tasks of the various health Institutions in the Italian integrated system for diagnosis, treatment, and surveillance of leprosy, we describe the management and outcomes of the leprosy case and of the patient's contacts. RESULTS: In April 2017, Multibacillary Lepromatose Leprosy was diagnosed in a 29-year-old Nigerian man who arrived in Rimini in July 2014 after a 2-year stay in Libya. The local Public Health Service implemented the epidemiological investigation and identified the patient's close contacts. The management of the case and the surveillance of the 13 identified contacts, 7 Italians and 6 migrants, highlighted some critical issues. The late diagnosis of the case, due to the lack of knowledge of exotic diseases by general practitioners and other health and social professionals, and the loss at follow up of the close contacts (5 out of 6 migrants), represented important obstacles to the full success of surveillance measures. CONCLUSIONS: Although in Italy there is a well codified system of notification and surveillance of leprosy, the recognising of cases and the tracing and follow up of contacts are made difficult by the particular conditions of the involved people. This represents a new challenge for the Italian Public Health Authorities which, in the current context of immigration, often uncontrolled, must know how to respond to the new needs, in close collaboration with the State Institutions responsible for registering migrants and those health and social professionals who could facilitate the access of foreign people to health services.

Axillary web syndrome assessment using a self-assessment questionnaire: a prospective cohort study.

BACKGROUND: Surgical procedure for breast cancer is not without its side effects and one such side effect is axillary web syndrome (AWS), characterized by palpable fibrotic-like cords in the operated arm. As physical evaluation is the only gold standard method used, our study aims to assess the incidence and early detection of AWS with a self-assessment questionnaire. METHODS: From July 2013 to July 2014, 370 breast cancer patients were enrolled. AWS incidence was 51.1%, with 94.1% onset in the first 4 weeks after surgery; 43.5% of the patients did not recover in the first 8 weeks. Univariate analysis showed that BMI (P < 0.001), age (P < 0.001), educational level (P = 0.01), and exercise frequency in the eighth week of follow-up (P < 0.001) were significantly associated with the AWS detection, and multivariate analyses confirmed that younger patients (age < 50) have significantly higher AWS detection (OR = 2.38 (95%CI 1.53, 3.71) and that BMI is associated with AWS, with normal weight patients (BMI ≤ 25) having a significantly greater AWS detection with an odds ratio of 2.11 (95%CI 1.33, 3.36). CONCLUSION: Our findings indicated that the incidence of AWS is high in breast cancer patients, particularly in the first month after surgery. Not all patients achieved recovery during our 8 week follow-up, suggesting that evaluation and treatment should be longer. Double AWS detection was found for patients who were younger (age < 50) and with normal weight.

Evaluation of the capillary electrophoresis method for measurement of immunoglobulin concentration in ewe colostrum.

Capillary electrophoresis (CE) is a technique routinely used in clinical laboratories that allows the separation and quantification of blood serum proteins in a rapid, precise, accurate, and inexpensive manner. Recently, CE has been proposed to separate and measure colostral proteins, but an evaluation of the agreement between CE and radial immunodiffusion (RID) method, currently used to quantify IgG in colostrum, is still lacking. The purpose of this study was to test the ability of a CE instrument, normally used in blood serum protein analysis, to realize the correct quantification of total Ig concentration in ewe colostrum, using RID assay as reference. Colostrum samples (n = 68) were collected from 35 multiparous Sarda ewes at first milking (n = 33) and at 24 h postpartum (n = 35). The mean ± standard deviation of IgG concentration measured by RID and whey colostrum total Ig concentration measured by CE were 54.76 ± 41.82 g/L and 54.70 ± 41.43 g/L, respectively. Lin's concordance correlation coefficient (r = 0.993; 95% confidence interval = 0.989 to 0.996) and linear regression analysis results (RID = 1.0022CE - 0.063; R2 = 0.986) showed an excellent agreement between these 2 methods. Bland-Altman analysis confirmed that CE method can be a suitable alternative to RID: the mean of the differences between CE and RID was -0.055 ± 4.95 g/L (95% confidence interval = -1.25 to 1.14 g/L) and the agreement limits were -9.75 to 9.60 g/L (low limit 95% confidence interval = -11.82 to -7.68 g/L; high limit 95% confidence interval = 7.57 to 11.72 g/L). In conclusion, the current study indicates that CE method may be a reliable tool for the quantification of the total Ig concentration in ewe colostrum.

High Blood Pressure States in Children, Adolescents, and Young Adults Associate Accelerated Vascular Aging, with a Higher Impact in Females' Arterial Properties.

The aims of the study were to determine (1) whether the presence of High blood pressure (HBP) states in the youth associate a steeper rate of age-related change in arterial geometrical and wall properties with respect to subjects with no previous cardiovascular risk factor (CRF) exposure, (2) in which parameters and in what magnitude, and (3) the existence of a gender-related difference in the impact of this condition on arterial properties. 300 individuals (mean/range: 15/4-29 years; 133 females) were included. Two groups were assembled: (1) Reference: nonprevious exposure to traditional CRF and (2) HBP: subjects with arterial hypertension and/or elevated blood pressure (BP) levels during the study. Additionally, HBP subjects were separated in BP-related subgroups. Measured parameters were (1) central (aortic) arterial BP and aortic pulse wave analysis parameters, (2) carotid and femoral artery local (pressure-strain elastic modulus) and regional (pulse wave velocity; PWV) stiffness, and (3) arterial diameters and carotid intima-media thickness (CIMT). Age-related changes in these parameters (absolute values and z-scores) were explored by obtaining simple linear regression models for each group. HBP presented a steeper rate of change (accelerated vascular aging; VA) for most of the parameters assessed, mainly in central (aortic) hemodynamics. VA increased as the HBP level got higher. Both males' and females' aging rates were affected by this condition, but females presented a more marked relative age-related increase with HBP exposure. HBP states in the youth gradually associate accelerated VA, with a progressive hemodynamic-structural-functional onset of damage, with females presenting a more marked relative HBP-associated arterial repercussion.

A New Surgical Model of Skeletal Muscle Injuries in Rats Reproduces Human Sports Lesions.

Skeletal muscle injuries are the most common sports-related injuries in sports medicine. In this work, we have generated a new surgically-induced skeletal muscle injury in rats, by using a biopsy needle, which could be easily reproduced and highly mimics skeletal muscle lesions detected in human athletes. By means of histology, immunofluorescence and MRI imaging, we corroborated that our model reproduced the necrosis, inflammation and regeneration processes observed in dystrophic mdx-mice, a model of spontaneous muscle injury, and realistically mimicked the muscle lesions observed in professional athletes. Surgically-injured rat skeletal muscles demonstrated the longitudinal process of muscle regeneration and fibrogenesis as stated by Myosin Heavy Chain developmental (MHCd) and collagen-I protein expression. MRI imaging analysis demonstrated that our muscle injury model reproduces the grade I-II type lesions detected in professional soccer players, including edema around the central tendon and the typically high signal feather shape along muscle fibers. A significant reduction of 30% in maximum tetanus force was also registered after 2 weeks of muscle injury. This new model represents an excellent approach to the study of the mechanisms of muscle injury and repair, and could open new avenues for developing innovative therapeutic approaches to skeletal muscle regeneration in sports medicine.

Lipid and non-lipid cardiovascular risk factors in postmenopausal type 2 diabetic women with and without coronary heart disease.

BACKGROUND: Coronary heart disease (CHD) is the leading cause of death in diabetic women. In addition to hyperglycemia, other factors may contribute to the excessive cardiovascular risk. AIM: In this study we evaluated common and emerging risk factors in a selected group of postmenopausal type 2 diabetic women with (n = 36) and without CHD (n = 59), not taking lipid-lowering medications. METHODS: Clinical and lifestyle data were collected, and metabolic and lipid profile, as well as fasting plasma levels of total homocysteine (tHcy), folate, vitamin B12, C-reactive protein (hsCRP), interleukin 6 (IL-6), and vascular cell adhesion molecule-1 (VCAM-1) were measured in all participants. RESULTS: Age, menopause and diabetes duration, family history for cardiovascular disease, prevalence of hypertension and current insulin use were greater in diabetic women with than without CHD (P < 0.05 for all comparisons). CHD women also showed higher levels of triglycerides, small dense LDL (sdLDL), remnant-like particle cholesterol, tHcy, and VCAM-1, and a lower creatinine clearance (P < 0.05 all). Conversely, the two groups were comparable for BMI, waist circumference, smoking habit, fasting plasma glucose, HbA1c, total cholesterol, low-density lipoprotein cholesterol (LDL-C), HDL cholesterol, folate, vitamin B12, hsCRP and IL-6 levels. At multivariate analysis, lower creatinine clearance (OR = 0.932, P = 0.017) and higher sdLDL serum concentration (OR = 1.224, P = 0.037) were the strongest risk factors associated with CHD in this population, whereas no significant association was noted with LDL-C. CONCLUSIONS: Our data suggest that beyond LDL-C, a lower creatinine clearance and more subtle alterations of LDL particles, together with a constellation of several well known and emerging cardiovascular risk factors, are stronger contributors to the high CHD risk of diabetic women.

[Herpes hepatic failure during the third quarter of pregnancy]. / Le cas clinique du mois. Hépatite herpétique au 3eme trimestre de la grossesse.

Acute liver diseases of pregnancy are common and usually transient and reversible. Given the number of different possible diagnoses, performing a large biological screening and a proper iconographic documentation is key. It makes sure no etiology fatal to the mother and her fetus is missed.

Neutral and acidic oligosaccharides in Holstein-Friesian colostrum during the first 3 days of lactation measured by high performance liquid chromatography on a microfluidic chip and time-of-flight mass spectrometry.

Oligosaccharides (OS) from bovine milk are a class of bioactive molecules that are receiving increasing commercial attention for their potential health benefits. In the present work we measured, comprehensively and systematically, free milk OS in the colostrum of 7 Holstein-Friesian cows during the first 3 d of lactation in 12-h intervals by HPLC-chip/time-of-flight mass spectrometry to determine the biological variation of free milk OS in early lactation. The high sensitivity and resolution of the analytical technique made it possible to monitor all OS species, thus providing a comprehensive and quantitative analysis of OS variations during colostrum production. This study confirmed that although sialyllactose is the major OS in bovine colostrum, several neutral OS species are present in significant abundance even at the third day of lactation. Furthermore, variation in terms of OS species and relative abundances of OS between cows suggest individual animal variation. These variations are likely due to genetic factors because environmental factors such as nutrition, lactation number, and accommodation were the same for all cows. This investigation revealed that colostrum milk from Holstein-Friesian cows is a rich source of neutral and acidic OS for the food and pharmaceutical industries.

Outcome of laparoscopic sacrocolpopexy with anterior and posterior mesh.

AIM: The assessment of the postoperative outcome following laparoscopic sacrocolpopexy using anterior and posterior mesh. MATERIAL AND METHODS: In the study were included one hundred and ten women (mean age 62 years with range from 34 to 78) who had laparoscopic sarcoplexy the period 2001-2005. They were contacted and completed postal questionnaires more than one year after surgery and had a follow up in the uro-gynaecology clinic. RESULTS: The median follow up was 21 months. Eighty-three of them (75.4%) answered the postal questionnaire. Good satisfaction was defined as complete disappearance of all genito-urinary symptoms. Moderate satisfaction was defined as a partial disappearance of symptoms, or de novo less annoying symptoms. Dissatisfaction was defined as no change in symptoms, and /or de novo important symptoms. The overall rate of good satisfaction was 74.7% (62/83), moderate satisfaction 15.6% (13/83), and only 9.6% (8/83) were not satisfied. There was no statistical difference among the six different groups (sacrocolpopexy only, plus TVT, plus TVT and hysterectomy, sacrocolpopexy and previous hysterectomy, sacrocolpopexy and TVT with previous hysterectomy), concerning the pre and post operative clinical signs and post operative symptoms. There was a statistically significant difference (p=0.038) regarding dissatisfaction and prolapse relapse between the group that had a previous total hysterectomy combining sacrocolpopexy with TVT and all other groups. The most frequent post operative symptoms were stress incontinence, dysuria and constipation. No severe complications and mesh erosion were observed, despite the two cases of mesh detachment. CONCLUSIONS: Laparoscopic double synthetic mesh sacrocolpopexy seems to be a safe and effective treatment of genitourinary prolapse, with good overall long term outcomes and benefits of the minimal access approach. The presence of the remaining cervix after subtotal hysterectomy, seems to enhance the results of laparoscopic sacrocolpopexy.. Further randomised studies are needed to confirm our results and to compare this method to open and/or vaginal approach.

18F-FDG positron autoradiography with a particle counting silicon pixel detector.

We report on tests of a room-temperature particle counting silicon pixel detector of the Medipix2 series as the detector unit of a positron autoradiography (AR) system, for samples labelled with (18)F-FDG radiopharmaceutical used in PET studies. The silicon detector (1.98 cm(2) sensitive area, 300 microm thick) has high intrinsic resolution (55 microm pitch) and works by counting all hits in a pixel above a certain energy threshold. The present work extends the detector characterization with (18)F-FDG of a previous paper. We analysed the system's linearity, dynamic range, sensitivity, background count rate, noise, and its imaging performance on biological samples. Tests have been performed in the laboratory with (18)F-FDG drops (37-37 000 Bq initial activity) and ex vivo in a rat injected with 88.8 MBq of (18)F-FDG. Particles interacting in the detector volume produced a hit in a cluster of pixels whose mean size was 4.3 pixels/event at 11 keV threshold and 2.2 pixels/event at 37 keV threshold. Results show a sensitivity for beta(+) of 0.377 cps Bq(-1), a dynamic range of at least five orders of magnitude and a lower detection limit of 0.0015 Bq mm(-2). Real-time (18)F-FDG positron AR images have been obtained in 500-1000 s exposure time of thin (10-20 microm) slices of a rat brain and compared with 20 h film autoradiography of adjacent slices. The analysis of the image contrast and signal-to-noise ratio in a rat brain slice indicated that Poisson noise-limited imaging can be approached in short (e.g. 100 s) exposures, with approximately 100 Bq slice activity, and that the silicon pixel detector produced a higher image quality than film-based AR.

Preparation and characterization of calcium phosphate biomaterials.

Calcium phosphate cement (CPC) samples have been prepared with a mixture of monocalciumphosphate monohydrate (MCPM) and calcium carbonate (CC) powders, in stechiometric moles ratio 1:2.5 to obtain a Ca/P ratio of about 1.67 typical of hydroxyapatite (HAp), with or without addition of HAp. All specimens are incubated at 30 degrees C in a steam saturated air environment for 3, 6 and 15 days respectively, afterwards dried and stored under nitrogen. The calcium phosphate samples have been characterized by X-ray diffraction (XRD), Vickers hardness test (HV), diametral compression (d.c.), strength compression, and porosity evaluation. MCPM/CC mixture has a 30% HAp final concentration and is characterized by higher porosity (amount 78%) and mechanical properties useful as filler in bone segments without high mechanical stress.

[Muscle regeneration following glycerol injection mimic that of mdx-mice degenerative-regenerative groups]. / La regeneración tras inyección intramuscular de glicerol remeda la de los grupos degenerativos-regenerativos del ratón mdx.

INTRODUCTION: In recent years several authors have proposed that muscle pathological changes in humans with dystrophin disorders and in mdx-mice are the result of regeneration attempts of this tissue subjected to constantly repeated bouts of necrosis. AIM: We compared morphological aspects of degenerative-regenerative groups observed in the mdx-mice with the regenerative phases that follow an intramuscular injection of 30 mL glycerol. MATERIALS AND METHODS: Gastrocnemius samples from mdx-mice, 1 to 5 months old and from glycerol injected C57Bl10/ScSn mice, 4 to 8 months old, were processed and analyzed in parallel. RESULTS: We were able to recognize degenerative-regenerative group stages in the mdx-mice that closely matched the anomalies found 1, 2, 3, 4-5, 7 and 21-28 days following the injection with glycerol. CONCLUSION. This study reinforces the hypothesis that muscle pathologic abnormalities of mdx-mice are the result of chronic muscle necrosis-regeneration. Morphologic mdx-mice degenerative-regenerative group staging will be a helpful tool for future investigation of muscle regeneration.

Correlation between cognitive impairment and the Rey auditory-verbal learning test in a population with Alzheimer disease.

Patients affected by Alzheimer disease (AD) need an accurate diagnosis, to the extent allowing us to find the best therapy or polytherapy, in order to take under control their cognitive impairment. In our Alzheimer Evaluation Units (from the Italian name abbreviated: UVA), the patients undergo a multidimensional evaluation, which can address us towards a proper diagnosis and of other weakening, or even dementia-related diseases. The patients are also subject to neuropsychometric and neuropsychological evaluations, allowing a more focused analysis on cognitive impairments. Among the tests, we use the Rey auditory-verbal learning test (RAVLT), evaluating the patient's verbal memory. A list of 15 words is read to each patient. N the first part of the test, the clinician repeats 5 times such a list. the patient is hen asked, at the end of every repetition, to tell all words he/she remembers. This part is useful to evaluate the immediate recall (IR) ability. The score, i.e., the total number of recalled words, ranges from 0 to 75. After 15 minutes, the delayed recall (DR) ability is evaluated: the patient is newly asked to repeat as many words as he can recall from the list. The score for this part ranges from 0 to 15 minutes. The score is corrected of rage and education, with a cut-off of 28.5 for IR and 4.7 for DR. We made a survey with the purpose of deciding if there was a correlation between cognitive impairment and verbal memory lack, whose deficiency appears earlier in AD. To this aim, we selected several patients with AD, diagnosed during the period between September 2002 and February 2003. We only considered those patients whose AD was not associated with other weakening diseases, and whose clinical dementia rating scale (CDR) score was between 0.5-2.0. A sample of 35 individuals (11 men and 24 women) could be obtained. A meaningful correlation was observed between CDR and IR (r = -0.725, p < 0.01), as well as between CDR and DR (r = -0.470; p < 0.05). Such a result confirms the importance of evaluating immediate and long-term memories, for the early diagnosis of AD, because it is the only symptom of clinically not yet diagnosed dementia, as proven also by other studies.

The management of psychogeriatric patient.

The exponential growth in the prevalence of cognitive impairment of old patients leads the physicians to deal with a larger incidence of behavioral disorders (such as excitement,aggressiveness), and psychotic symptoms (such as delirium and visual hallucinations). The presence of psychotic troubles in dementia causes a remarkable distress to caregivers and involves higher difficulties in the patient management. The estimates of such troubles range between 15 and 75 %. Geriatric assessment and the management of behavioral troubles require a prompt evaluation of all their possible causes. As a matter of fact, their appearance often reveals a physical disturbance (pain, fever, etc.), or adverse environmental conditions, or it could also be a consequence of a multiple drug therapy. For this reason,the use of antipsychotics should always be preceded by an accurate clinical diagnosis.Anxiolytic, anti-depressive, anti-convulsive and anti-psychotic drugs are among the therapeutic strategies for the management of the psychogeriatric patient. Atypical antipsychotics seem to be able to decrease the psychotic symptoms, with low levels of therapeutic failure. They also reduce extrapyramidal effects and the growth of prolactine hormone, which is quite useful when dealing with very old patients. Risperidone and olanzapine are two atypical anti-psychotics, which already proved to be adequate and well tolerated during the treatment of schizophrenia and of acute maniacal disorders. Our experience, with a population of patients followed by our Alzheimer Evaluation Unit (AEU), confirms that a low dose of olanzapine (5mg/day) and risperidone (0.5-1.0 mg/day) are effective in lowering behavioral disturbances, and psychotic symptoms due to dementia. Even in the long run,low doses of these drugs are still well tolerated. Higher levels of risperidone (> 1 mg/die)often caused extra-pyramidal symptoms such as rigidity and dyskinesia, whereas higher levels of olanzapine (> 5 mg/day) lead to an exceeding sedation. The management of behavioral disturbances is one of the most important goals in the global treatment of patients affected by dementia, to the extent of improving the quality of life. Atypical antipsychotics are preferable compared to old-generation drugs, therefore, they are the key therapeutic strategy we cannot renounce.

La regeneración tras una inyección intramuscular de glicerol remeda la de los grupos degenerativos-regenerativos del ratón mdx / Muscle regeneration following glycerol injection mimic that of mdx-mice degenerative-regenerative groups

Introducción. En los últimos años, diversos autores han sugerido que los cambios patológicos musculares en los humanos con distrofinopatías y en los ratones mdx son el resultado de los intentos de regeneración realizados por este tejido sujeto a repetidas lesiones por necrosis. Objetivo. Se compararon los aspectos morfológicos de los grupos degenerativos-regenerativos observados en los ratones mdx y las fases regenerativas que se producen después de una inyección intramuscular de 30 µL de glicerol. Materiales y métodos. Se trataron y se analizaron unas muestras de gastrocnemio tomadas de ratones mdx, de 1 a 5 meses de edad, y de ratones C57Bl10/ScSn en los cuales se había inyectado glicerol, de 4 a 8 meses de edad. Resultados. Se reconocieron estadios en el grupo degenerativo-regenerativo en los ratones mdx que tenían una estrecha correspondencia con las anomalías halladas 1, 2, 3, 4-5, 7 y 21-28 días después de la inyección con glicerol. Conclusión. Este estudio corrobora la hipótesis de que las anomalías patológicas musculares de los ratones mdx son el resultado del proceso crónico de necrosis-regeneración muscular. La determinación del estadio morfológico del grupo degenerativoregenerativo de los ratones mdx será un instrumento valioso en futuras investigaciones (AU)

Introduction. In recent years several authors have proposed that muscle pathological changes in humans with dystrophin disorders and in mdx-mice are the result of regeneration attempts of this tissue subjected to constantly repeated bouts of necrosis. Aim. We compared morphological aspects of degenerative-regenerative groups observed in the mdx-mice with the regenerative phases that follow an intramuscular injection of 30µL glycerol. Materials and methods. Gastrocnemius samples from mdx-mice, 1 to 5 months old and from glycerol injected C57Bl10/ScSn mice, 4 to 8 months old, were processed and analyzed in parallel. Results. We were able to recognize degenerative-regenerative group stages in the mdx-mice that closely matched the anomalies found 1, 2, 3, 4-5, 7 and 21-28 days following the injection with glycerol. Conclusion. This study reinforces the hypothesis that muscle pathologic abnormalities of mdx-mice are the result of chronic muscle necrosisregeneration. Morphologic mdx-mice degenerative-regenerative group staging will be a helpful tool for future investigation of muscle regeneration (AU)

Expression of Sulfolobus solfataricus alpha-glucosidase in Lactococcus lactis.

The industrial potential to use extreme thermophilic microorganisms and their enzymes lies in applications in which the temperature cannot be adjusted (cooled) at will. The production of enzymes from wild-type thermophiles is very low, therefore, for industrial applications, it is necessary to use recombinant microorganisms. In this paper, the cloning of a heat-stable alpha-glucosidase from Sulfolobus solfataricus using lactic acid bacteria as expression system is reported. The extremophilic alpha-glucosidase was cloned in Lactococcus lactis and correctly folded despite being expressed at a lower temperature. The recombinant cells were assayed for enzyme residual activity at 75 degrees C in order to analyze the direct use of whole cells as biocatalysts. Maximum activity corresponded to 40 U/l in static cultures. The protein yield was further improved by optimizing fermentation and reached 600 U/l in batch mode. Microfiltration led to an even higher enzyme production of 850 U/l as a result of increased biomass. The overall production of alpha-glucosidase using the engineered L. lactis strain in microfiltration fermentation is 1,000-fold higher than obtained using the wild-type.

Adenovirus-mediated wt-p16 reintroduction induces cell cycle arrest or apoptosis in pancreatic cancer.

Pancreatic cancer has long carried poor prognosis. The development of new therapeutic approaches is particularly urgent. Inactivation of the tumor-suppressor gene p16(INK4a/CDKN2), a specific inhibitor of the cyclin-dependent kinases CDK4 and CDK6, is the most common genetic alteration in human pancreatic cancer, making it an ideal target for gene replacement. Here we transfected tumor cells using a recombinant adenovirus containing the wt-p16 cDNA (Ad5RSV-p16). The overexpression of p16 decreased cell proliferation in all four human pancreatic tumor cell lines (NP-9, NP-18, NP-29, and NP-31). However, G1 arrest and senescence were observed in only three. In contrast, the fourth (NP-18) showed a significant increase in apoptosis. This differential behavior may be related to the differences found in the expression level of E2F-1. Experiments on subcutaneous pancreatic xenografts demonstrated the effectiveness of p16 in the inhibition of pancreatic tumor growth in vivo. Taken together, our results indicate that approaches involving p16 replacement are promising in pancreatic cancer treatment.

DAG accumulation from saturated fatty acids desensitizes insulin stimulation of glucose uptake in muscle cells.

The increased availability of saturated lipids has been correlated with development of insulin resistance, although the basis for this impairment is not defined. This work examined the interaction of saturated and unsaturated fatty acids (FA) with insulin stimulation of glucose uptake and its relation to the FA incorporation into different lipid pools in cultured human muscle. It is shown that basal or insulin-stimulated 2-deoxyglucose uptake was unaltered in cells preincubated with oleate, whereas basal glucose uptake was increased and insulin response was impaired in palmitate- and stearate-loaded cells. Analysis of the incorporation of FA into different lipid pools showed that palmitate, stearate, and oleate were similarly incorporated into phospholipids (PL) and did not modify the FA profile. In contrast, differences were observed in the total incorporation of FA into triacylglycerides (TAG): unsaturated FA were readily diverted toward TAG, whereas saturated FA could accumulate as diacylglycerol (DAG). Treatment with palmitate increased the activity of membrane-associated protein kinase C, whereas oleate had no effect. Mixture of palmitate with oleate diverted the saturated FA toward TAG and abolished its effect on glucose uptake. In conclusion, our data indicate that saturated FA-promoted changes in basal glucose uptake and insulin response were not correlated to a modification of the FA profile in PL or TAG accumulation. In contrast, these changes were related to saturated FA being accumulated as DAG and activating protein kinase C. Therefore, our results suggest that accumulation of DAG may be a molecular link between an increased availability of saturated FA and the induction of insulin resistance.

Changes in myocardial cytoskeletal intermediate filaments and myocyte contractile dysfunction in dilated cardiomyopathy: an in vivo study in humans.

AIM: To investigate in vivo the intermediate cytoskeletal filaments desmin and vimentin in myocardial tissues from patients with dilated cardiomyopathy, and to determine whether alterations in these proteins are associated with impaired contractility. METHODS: Endomyocardial biopsies were performed in 12 patients with dilated cardiomyopathy and in 12 controls (six women with breast cancer before anthracycline chemotherapy and six male donors for heart transplantation). Biopsy specimens were analysed by light microscopy and immunochemistry (desmin, vimentin). Myocyte contractile protein function was evaluated by the actin-myosin in vitro motility assay. Left ventricular ejection fraction was assessed by echocardiography and radionuclide ventriculography. RESULTS: Patients with dilated cardiomyopathy had a greater cardiomyocyte diameter than controls (p < 0.01). The increase in cell size was associated with a reduction in contractile function, as assessed by actin-myosin motility (r = -0.643; p < 0.01). Quantitative immunochemistry showed increased desmin and vimentin contents (p < 0.01), and the desmin distribution was disturbed in cardiomyopathy. There was a linear relation between desmin distribution and actin-myosin sliding in vitro (r = 0.853; p < 0.01) and an inverse correlation between desmin content and ejection fraction (r = -0.773; p < 0.02). Negative correlations were also found between myocardial vimentin content and the actin-myosin sliding rate (r = -0.74; p < 0.02) and left ventricular ejection fraction (r = -0.68; p < 0.01). CONCLUSIONS: Compared with normal individuals, the myocardial tissue of patients with dilated cardiomyopathy shows alterations of cytoskeletal intermediate filament distribution and content associated with reduced myocyte contraction.

Computer-assisted analysis of methylation status of individual interphase nuclei in human cultured cells.

This paper demonstrates that (a) differences in the methylation levels of interphase nuclei can be measured on a cell-by-cell basis, (b) the binding sites of beta-satellite DNA and 5-methylcytosine (5MeC)-rich regions can be localised in interphase nuclei and metaphase chromosomes by sequential in situ hybridization and indirect immunolabelling, and (c) quantitative differences in the relative extensions of beta-satellite DNA and anti-5MeC antibody binding areas can also be measured. This goal was achieved by indirect immunolabelling by anti-5MeC antibodies (Reynaud et al.: Cancer Lett. 61:255-262, 1991) of control and 5-azacytidine-treated human cell cultures. A quantitative analysis of the number, total, and mean areas of labelled heterochromatic regions and the optical densities of euchromatin and heterochromatin was performed for the cells on microscope slides. Dedicated software was used to select and measure the areas of cytological interest. In additional experiments, DAPI-stained slides from control cultures were sequentially treated by in situ hybridization with beta-satellite DNA probe and indirect immunofluorescent labelling with anti-5MeC antibodies. Fluorescent signals of probe and antibodies were pseudocoloured and merged on digital images. The relative locations of probe- and antibody-positive areas were analysed on metaphases and nuclei, and their extensions were quantified in interphase nuclei. Our results show that (a) our analysis can successfully detect different levels of DNA methylation within individual nuclei, (b) in metaphase chromosomes the antibody binding sites are mostly coincident with the hybridisation sites, and (c) in interphase nuclei a quite different picture is consistently observed.