Massimo Locatelli

In vivo evaluation of timing, degree, and distribution of bacterial translocation following experimental small bowel transplantation

These studies were designed to evaluate the correlation between morphologic and functional changes after heterotopic auxiliary small bowel isograft with systemic venous drainage and two ostomies in 20 Lewis rats. Morphologic damage of the graft was scored by full-thickness biopsies before surgery and 1, 3, 5, and 7 days after transplant. Functional evaluation of the graft was done, at the same time points, by urinary excretion of lactulose and mannitol injected in the proximal ostomy. The intestinal permeability was also studied by injecting Escherichia coli labeled with indium-111 oxine in the proximal ostomy. Translocation of radiolabeled bacteria was quantitated in extraintestinal tissues by radionuclide counts and number of viable organisms and in vivo by scinti-graphic imaging. One day after transplant, significant graft damage (score 17.2±4.2) was observed when compared with the pretransplant value (7.3±2.6). The degree of tissue injury was similar on days 3 (15.8±3.5) and 5 (16.1 ±3.9) after transplant and remained high on day 7 (11.8±2.8). The lactulose to mannitol ratio showed a significantly increased permeability on day 1 (17.5) versus pretransplant values (2.6), remained high on day 3 (8.6), and returned to normal values on day 5 (2.8). Translocation of bacteria to distant organs, as measured by both radionuclide counts and number of viable organisms, was strikingly enhanced on day 1 after transplantation, compared with control animals, but returned to the pretransplant value on day 3. A good qualitative and quantitative correlation was observed between radionuclide counts in the extraintestinal organs and in vivo images obtained by scintigraphic scanning. In conclusion, in this model, timing and degree of bacterial translocation do not seem to correlate well, with the exception of the acute posttransplantation phase, with morphologic and perme ability changes of the graft. Evaluation of translocation by scintigraphic imaging appears a suitable approach to study in vivo the kinetics and distribution of this process.

Beneficial effects of beta-blockers on left ventricular function and cellular energy reserve in patients with heart failure

Beta‐blockers have been shown to improve left ventricular (LV) function in patients with heart failure. The aim of this study is to non‐invasively assess, by means of in vivo 31P‐magnetic resonance spectroscopy (31P‐MRS), the effects of beta‐blockers on LV cardiac phosphocreatine and adenosine triphosphate (PCr/ATP) ratio in patients with heart failure. Ten heart failure patients on full medical therapy were beta‐blocked by either carvedilol or bisoprolol. Before and after 3 months of treatment, exercise testing, 2D echocardiography, MRS, New York Heart Association (NYHA) class, ejection fraction (EF), maximal rate–pressure product and exercise metabolic equivalent system (METS) were evaluated. Relative concentrations of PCr and ATP were determined by cardiac 31P‐MRS. After beta‐blockade, NYHA class decreased (from 2.2 ± 0.54 to 1.9 ± 0.52, P = 0.05), whereas EF (from 33 ± 7 to 44 ± 6%, P = 0.0009) and METS (from 6.74 ± 2.12 to 8.03 ± 2.39, P = 0.01) increased. Accordingly, the mean cardiac PCr/ATP ratio increased by 33% (from 1.48 ± 0.22 to 1.81 ± 0.48, P = 0.03). Beta‐blockade‐induced symptomatic and functional improvement in patients with heart failure is associated to increased PCr/ATP ratio, indicating preservation of myocardial high‐energy phosphate levels.

Comparison of the Hematological Profile of Elite Road Cyclists during the 2010 and 2012 GiroBio Ten-Day Stage Races and Relationships with Final Ranking

Cycling stage races are strenuous endurance events during which exercise-induced variations in hematological parameters are consistently observed. However, specific literature on such changes is scarce and published data have been derived from small samples of athletes. The aims of this study were: (1) to determine the hematological response to middle-term strenuous endurance; and (2) to determine whether a relationship exists between the athlete-specific hematological profile and final placement in a cycling stage race. The study population was male professional cyclists (n = 253) competing in the 2010 (n = 144) and 2012 (n = 109) GiroBio 10-day stage races. Blood draws taken before the start of the race, at mid-race, and at end-race were performed in strict compliance with academic and anti-doping pre-analytical warnings. Blood chemistry included white blood cell, red blood cell, hemoglobin concentration, hematocrit, mean corpuscular volume (MCV), mean hemoglobin content (MCH), mean corpuscular hemoglobin content (MCHC), platelets, and reticulocyte relative and absolute counts. Compared to baseline values, erythrocyte, hemoglobin, hematocrit, MCHC, platelet and reticulocyte counts were all consistently lower at mid-race, but returned to normal by race-end, while leukocytes were increased in the final phase. MCV increased during both events. MCH increased in the first part to then return to baseline in the 2012 race. The calculated OFF-score consistently decreased in the first half of the race before increasing, but remained lower than the baseline value. The trends of variation in hematological parameters were substantially similar in both events. There was an inverse, albeit weak, relationship between placement and erythrocyte, platelet, hemoglobin, hematocrit and OFF-score values in the 2010, but not in the 2012 race. In conclusion, the data confirm that, in this large series of elite road cyclists, the strenuous effort a rider sustains during a stage race induces appreciable changes in the hematological profile.

Clinical significance of the number of oligoclonal bands in patients with clinically isolated syndromes

CSF oligoclonal bands (OCBs) in patients with clinically isolated syndromes (CIS) are a risk factor for clinically definite multiple sclerosis (CDMS). We aimed to address the relevance of the number of OCBs in the prognosis of CIS patients. 219 CIS patients were included in the study, and 42% of them developed the disease during follow-up (median: 5.04 years). Patients with a high number of CSF OCBs (third quartile, 8-12 OCBs) had 2.5-fold increase in CDMS risk, while no further increase in the HR of disease was observed for patients with more than 12 OCBs. The results did not change after adjustment for additional correlates of CDMS development. This association may be due to the epitope-spreading phenomenon and may reflect the stage of the disease at the time of the examination.

Increased low-grade inflammation is associated with lack of functional response to carvedilol in patients with systolic heart failure.

Background The aim of this study was to evaluate, according to functional response, the neuroendocrine and inflammatory status in patients with chronic heart failure before and after therapy with carvedilol. Methods and results Serum tumor necrosis factor-&agr; (TNF-&agr;) soluble receptors (sTNF-R1 and sTNF-R2), interleukin (IL)-10 and IL-18, chromogranin A (CgA) and brain natriuretic peptide (pro-BNP) were measured in 37 New York Heart Association class II to IV heart failure patients, at baseline and after 6 months of therapy with carvedilol. Patients were divided in two groups according to whether, following carvedilol, left-ventricular ejection fraction (LVEF) had increased by at least 5% (17 patients) or not (20 patients). Baseline LVEF was higher in nonresponders (38 ± 5 vs. 31 ± 7%, P = 0.002). In responders, LVEF increased from 31 ± 7 to 51 ± 7% (P < 0.0001), whereas in nonresponders it decreased from 38 ± 5 to 33 ± 7%, (P = 0.02). sTNF-R1 (P = 0.019) and sTNF-R2 (P = 0.025) increased in nonresponders, whereas they did not change in responders. After carvedilol, IL-10 was significantly higher in responders (P = 0.03). Conversely, no significant IL-18 and CgA changes were observed in either group. CgA was not significantly different between groups at baseline and after carvedilol in either group, whereas pro-BNP significantly increased in nonresponders (from 438 ± 582 to 1324 ± 1664 pg/ml, P = 0.04) and decreased in responders (from 848 ± 1221 to 420 ± 530 pg/ml, P = 0.08). Conclusion Increased inflammatory activation observed only in heart failure patients not improving left-ventricular function after carvedilol may indicate that inflammation, either as a direct cause or as a consequence, is associated with progressive ventricular dysfunction.

Tumor-associated trypsin inhibitor in induced and acquired immunodeficiency. Studies on transplanted and HIV-infected patients.

A new tumor marker, tumor-associated trypsin inhibitor (TATI), was studied in 5 patients who received successful kidney or pancreas grafts and in 30 subjects with antibodies against human immunodeficiency virus. Serum TATI concentrations were very high during the four first days after transplantation. Thereafter the serum levels decreased when the peptide was eliminated through the kidney. Consequently, the urine values were very high. The TATI concentrations of HIV positive subjects were compared with serum levels of HIV antigen and antibody, by Western blotting and determination of peripheral T-lymphocyte subpopulations. The occurrence of high concentrations of TATI in some HIV positive subjects and especially in AIDS patients, suggests that TATI could be useful in exploring physiopathological aspects of severe immunodeficiencies even if TATI levels were not correlated with the commonly used markers of the immune system status. The increased levels of TATI in immunological disorders suggests its possible use in assessing the immune response against cancer.

Free Light Chains and Intrathecal B Cells Activity in Multiple Sclerosis: A Prospective Study and Meta-Analysis.

Background. The presence of CSF oligoclonal bands (OBs) is an independent prognostic factor for multiple sclerosis (MS), but the difficulties in the standardization of the test and the interlaboratory variation in reporting have contributed to its limited use in the diagnosis of the disease. Standard nephelometric assays to measure free light chains (FLC) levels have been recently developed and the test may improve the detection of intrathecal B cells activity. Methods. The presence of OBs, kappa and lambda FLC levels, and standard indices of intrathecal inflammation were assessed in 100 consecutive patients, including patients with MS, clinically isolated syndromes (CIS), other inflammatory diseases of the CNS, and other noninflammatory diseases. Results. Both KFLC and LFLC correlated strongly with the presence of OCBs and with all common tests for intrathecal inflammation (p < 0.001 for all comparisons). KFLC and LFLC were significantly different in patients with MS and CIS compared to the other groups (p < 0.001 and p < 0.001, resp.) and had a better diagnostic accuracy than all the other tests (area under the curve 82.3 % for KFLC index and 79.3 % for LFLC index). Conclusion. Nephelometric assays for KFLC in CSF reliably detect intrathecal immunoglobulin synthesis and discriminate MS patients.

Novel bone metabolism-associated hormones: the importance of the pre-analytical phase for understanding their physiological roles

The endocrine function of bone is now a recognized feature of this tissue. Bone-derived hormones that modulate whole-body homeostasis, are being discovered as for the effects on bone of novel and classic hormones produced by other tissues become known. Often, however, the data regarding these last generation bone-derived or bone-targeting hormones do not give about a clear picture of their physiological roles or concentration ranges. A certain degree of uncertainty could stem from differences in the pre-analytical management of biological samples. The pre-analytical phase comprises a series of decisions and actions (i.e., choice of sample matrix, methods of collection, transportation, treatment and storage) preceding analysis. Errors arising in this phase will inevitably be carried over to the analytical phase where they can reduce the measurement accuracy, ultimately, leading discrepant results. While the pre-analytical phase is all important, in routine laboratory medicine, it is often not given due consideration in research and clinical trials. This is particularly true for novel molecules, such as the hormones regulating the endocrine function of bone. In this review we discuss the importance of the pre-analytical variables affecting the measurement of last generation bone-associated hormones and describe their, often debated and rarely clear physiological roles.

Alcohol and illicit drugs in drivers involved in road traffic crashes in the Milan area. A comparison with normal traffic reveals the possible inadequacy of current cut-off limits

BACKGROUND Driving under the influence of alcohol and/or illicit drugs in Italy is regulated by the articles 186 and 187 of the National Street Code. Epidemiological studies on drivers involved in road traffic crashes (RTC) provide useful information about the use/abuse of these substances in the general population. Comparison with case control studies may reveal important information like the cut-off limits adequacy. METHODS Data from 1587 blood tests for alcohol and 1258 blood tests for illicit drugs on drivers involved in RTC around Milan between 2012 and 2016, were analyzed and compared with a published random survey (DRUID) from the European Community. RESULTS Our data from RTC-involved drivers show that alcohol abuse is not age-related whereas illicit drugs are more common in young people. Cannabinoids are frequent among younger drivers (median age 27) whereas cocaine is more often detected in adults (median age 34). The calculated odds ratio after comparison with the DRUID survey shows that a blood alcohol concentration below the legal limit does not represent a risk factor in having a car accident whereas concentrations of cocaine and cannabinoids within the legal limits are associated with being involved in a car accident. CONCLUSIONS Despite authority efforts, the abuse of alcohol and illicit drugs is still common in young drivers. We suspect that the cut-off limits for cannabinoids and cocaine and/or the pre-analytical procedures for these substances are inadequate. We suggest a better standardization of the procedure by shortening the time interval between the request for investigation and blood collection and propose the adoption of more stringent cut-off limits.

Toxicological investigation in blood samples from suspected impaired driving cases in the Milan area: Possible loss of evidence due to late blood sampling

Driving under the influence of illicit drugs (DUID) represents a significant menace to public safety and is therefore sanctioned with severe fines and penalties such as driving disqualification or even arrest in case the accident has caused serious injury or death. In Italy, DUID is regulated by the article 187 of the National Street Code, however, the list of the substances to be searched and their threshold concentrations are left to the 20 Italian regional authorities. A further lack of legislative standardization concerns the type of detection methods and moreover the time gap between the car accident and blood sampling. This interval can be as high as 5h, enough to significantly reduce the concentration of drugs with fast pharmacokinetic. By analyzing 1258 blood tests performed on drivers involved in road traffic crashes in the Milan area between 2012 and 2016 we show that approximately 75% of such drivers who tested positive for THC and 15% of the drivers who tested positive for cocaine are at risk of misjudgment. Considering the severe sanctions associated with DUID, we emphasize the urgency of introducing a corrective factor that takes into account the time elapsed between the accident and blood sampling in order to avoid unfair treatment, including the unjust application of sanctions.