Comparative Evaluation of Ligation-Mediated PCR and Spoligotyping as Screening Methods for Genotyping of Mycobacterium tuberculosis Strains

Spoligotyping has been suggested as a screening test in multistep genotyping of Mycobacterium tuberculosis strains. Relying on restriction fragment length polymorphism (RFLP) analysis with IS6110 (IS6110 RFLP analysis) as a "gold standard," we performed a comparative evaluation of spoligotyping and ligation-mediated PCR (LMPCR), a recently described PCR-based typing method, as rapid screening tests for fingerprinting of 158 M. tuberculosis strains collected in Verona, Italy. LMPCR seemed to be comparable to spoligotyping in terms both of feasibility with rapidly extracted DNA and of generation of software-analyzable images. Moreover, LMPCR grouped considerably fewer strains than spoligotyping (38 versus 67%) and was found to reduce the cluster overestimation rate (26.3 versus 58%) and to give a better discriminatory index (0.992 versus 0.970) compared to spoligotyping. In our geographical region, where there was no evidence of clustered strains carrying fewer than six IS6110 copies, LMPCR was found to be more discriminatory than spoligotyping. We also evaluated two models of three-step typing strategies, involving the use of spoligotyping and LMPCR as screening methods and IS6110 RFLP analysis as a further supporting test. LMPCR proved to be a more effective first-step test than spoligotyping, significantly reducing the need for subtyping. LMPCR should be considered an alternative to spoligotyping as a rapid screening method for M. tuberculosis fingerprinting, particularly in areas with a low prevalence of M. tuberculosis strains carrying few copies of IS6110.     

Pretransplant serum FT3 levels in kidney graft recipients are useful for identifying patients with higher risk for graft failure

Objective End‐stage renal disease (ESRD) is a condition associated with thyroid disturbances both in function and morphology. Recent studies demonstrated that serum free triiodothyronine 3 (FT3) levels are negatively correlated with serum markers of inflammation and endothelial activation in patients with ESRD. However, no previous research evaluated serum thyroid function parameters in relation to kidney graft outcome, as we aim to do so in this study. Design Serum FT3, free thyroxine 4 (FT4) and TSH levels were measured before transplantation in 196 kidney graft recipients. Results The graft survival rate at 5 years for all patients was 92·3%. Kidney graft recipients with normally functioning grafts showed serum pretransplant thyroid parameters similar to patients who experienced graft failure. Life‐time analysis was performed after stratification of patients according to pretransplant serum FT3 levels < 3·1 pmol/l or > 3·1 pmol/l. A significantly different 5‐year death‐censored graft survival rate (93·9%vs. 76·5% for patients with normal or low FT3 levels, respectively; P < 0·01) and similar survival rate (death of patients with functioning grafts) (21·1%vs. 5·9%; P = 0·288) were observed. No similar feature was found for FT4 or TSH, suggesting that the effect is not related to hypothyroidism but rather dependent upon inappropriately low FT3 levels. Pretransplant serum FT3 levels were similar in patients who experienced early acute rejections as compared with nonrejector patients. Conclusions The results of this study demonstrate that among patients with ESRD undergoing kidney transplantation, those displaying lower pretransplant serum FT3 levels are at higher risk for subsequent graft failure. The demonstration of a predictive value of serum FT3 levels for graft survival suggests that measurement of pretransplant serum FT3 levels might represent a clinically useful parameter to identify patients with increased risk for graft failure.     

White matter hyperintensities and their associations with suicidality in patients with major affective disorders

INTRODUCTION: A large body of evidence suggests that predisposition to suicide, an important public health problem, is mediated to a certain extent by neurobiological factors. The objective of this cross-sectional study was to compare the prevalence of white matter hyperintensities (WMH) in patients with major affective disorders with and without histories of suicide attempts. METHODS: T2-weighted magnetic resonance images (MRI) of 65 psychiatric inpatients with major depressive disorder or bipolar disorder were rated for the presence of WMH. Diagnoses, presence or absence of suicide risk and substance abuse were determined by the Mini International Neuropsychiatric Interview (MINI). Medical charts were reviewed to ascertain history of suicide attempt and basic clinical variables. Fisher's Exact Tests and logistic regression modeling were used to test the association between WMH and suicidality. Suicidal patients and controls were not matched for demographic variables and exposure to some risk factors. RESULTS: Bivariate analysis showed that the prevalence of WMH was significantly higher in subjects with past suicide attempts (Fisher's Exact Test, p = 0.01) and other clinical indicators of elevated suicide risk. Logistic regression analyses controlling for age, sex, and several clinical risk factors supported this finding (odds ratio = 4.7; 95% confidence interval: 1.4, 16.1). CONCLUSIONS: The increased prevalence of WMH in adults with major affective disorders and a history of suicide attempt, compared to similar patients without such a history, is consistent with previous findings in depressed children, youth and young adults. However, the association between WMH and suicidality holds true for both, depressed and bipolar patients. Our results suggest that WMH in patients with major affective disorders might be useful biological markers of suicidality.     

Fragile X premutation with atypical symptoms at onset

OBJECTIVE: To evaluate the presence of carriers of the fragile X premutation among male patients with sporadic ataxia without expansion into known spinocerebellar ataxia genes. DESIGN: Clinical and genetic examinations were performed on patients with sporadic pure ataxia and patients with ataxia associated with extracerebellar features such as pyramidal and extrapyramidal signs, dementia, or peripheral neuropathy. SETTING: University department of neurology. PATIENTS: One hundred forty-two Italian men with sporadic ataxia with onset at age 30 to 84 years. INTERVENTIONS: The CGG repeat size of the FMR1 gene was evaluated with fluorescent polymerase chain reaction. Premutated allele lengths were confirmed with Southern blot analysis. RESULTS: FMR1 premutation alleles with a repeat number greater than 55 were detected in 3 probands (2.1%) from a total of 142 male subjects initially referred to our university medical center for evaluation of sporadic ataxia. Two patients had typical fragile X syndrome with associated tremor or ataxia, and the third patient had spastic paraparesis without clear symptoms of cerebellar ataxia and without the common signs seen at magnetic resonance imaging. CONCLUSIONS: Genetic analysis of the FMR1 gene could provide a reliable diagnostic tool for the definitive diagnosis of late-onset ataxias. Additional studies are needed to clarify the importance of premutation screening in patients with movement disorders or other associated atypical features at onset, such as paraparesis.     

Acute pancreatitis in elderly patients: a single-center retrospective evaluation of clinical outcomes

Introduction: Acute pancreatitis (AP) incidence in the elderly population has increased in the last years. However, the role of age as influencing factor on the AP clinical course is still debated.

Methods: We reviewed clinical records of consecutive patients admitted with diagnosis of AP. Patients were divided in elderly (≥65 years) and non-elderly (<65 years). Primary endpoint was comparison of overall mortality. Secondary endpoint included ICU admission, in-hospital length of stay (LOS) and surgical procedures.

Results: We enrolled 352 elderly and 532 non-elderly patients. A higher mortality rate (7.4% vs 1.9%; p?<?.001), ICU admission rate (18.9% vs 6.3%; p?<?.001) and prolonged length of hospital stay (9 (6–14) vs 7 (5–11.7) days; p?=?.01) were registered in the ≥65 years group. Multivariate analysis identified age (OR: 3.5; 95% CI:1.645–7.555; p?=?.001), a higher Ranson score at admission (OR: 5.52; 95% CI:1.11–27.41; p<.001) and necrotic pancreatitis (OR: 8.6; 95% CI:2.46–30.27; p?=?.001) as independent predictors of mortality. Conversely age and necrotic pancreatitis were independent risk factors for higher LOS and ICU admission.

Conclusions: Patients with AP and age ≥65 years have a higher mortality, ICU admission and prolonged LOS. Early recognition and prompt treatment are key elements to improve outcomes in this population.     

Prognostic significance of cyclooxygenase-2 pathway and angiogenesis in head and neck squamous cell carcinoma

Prostaglandins play a critical role in tumor development and growth by regulating numerous biologic processes, including tumor angiogenesis, with clear prognostic and therapeutic implications. The aim of this study was to investigate the prognostic relevance of cyclooxygenase-2 (COX-2) pathway activation in head and neck squamous cell carcinoma (HNSCC). COX-2 activity was analyzed in 52 consecutive patients by assessing protein expression and prostaglandin E(2) (PgE(2)) levels and was then correlated to vascular endothelial growth factor (VEGF) expression and tumor angiogenesis. We evaluated the prognostic impact of these parameters by Kaplan-Meier and Cox survival analysis. COX-2 expression by tumor cells was closely correlated to VEGF expression and to tumor vascularization. According to Kaplan-Meier analysis, patients with COX-2 tumor overexpression and with higher PgE(2) tumor levels had significantly shorter overall survival estimates (P = 0.022 and P = 0.033, respectively). Analogously, patients with more-vascularized tumors had worse survival than those with less-vascularized cancers (P = 0.032). Cox multivariate analysis demonstrated that the most significant prognostic factors were presence of lymph node metastasis, tumor vascularization, COX-2 protein expression, and PgE(2) tumor levels. This study demonstrates a close correlation between COX-2 pathway, VEGF expression, and tumor angiogenesis in HNSCC. In addition, COX-2 overexpression and higher tumor vascularization appear to predict a shorter survival in patients with head and neck cancer.     

Psychopathological traits and 5-HT2A receptor promoter polymorphism (-1438 G/A) in patients suffering from Anorexia Nervosa and Bulimia Nervosa

Various studies have evaluated the possible role of the -1438G/A polymorphism within the 5-HT2A receptor gene in the susceptibility to Eating Disorders (EDs). One hundred and forty-eight ED patients (EDp) and 89 control subjects were interviewed by means of the Eating Disorder Examination (EDE) and analyzed for distribution of the -1438G/A polymorphism. Patients with the AA genotype suffering from Anorexia Nervosa and Bulimia Nervosa showed higher Weight and Shape Concern (P = 0.003 and P = 0.010, respectively) scores and greater overall severity of the ED psychopathology (EDE total score) (P = 0.012). The obtained preliminary data suggest the use of dimensional psychopathological measures in ED genetic studies.     

Kimura’s disease: case report of an Italian young male and response to oral cyclosporine A in an 8 years follow-up

Kimura’s disease is a benign chronic inflammatory disease, common in Asian males and rare in Western people. Clinically, Kimura’s disease is characterized by subcutaneous nodular lesions, usually localised in head and neck, often associated with regional lymphadenopathy. Peripheral blood eosinophilia and elevated serum IgE are often observed. We report a case of a 40-year-old Italian patient presenting with nodular subcutaneous lesions and peripheral eosinophilia. Based on clinical, histopathological and laboratory findings, a diagnosis of Kimura’s disease was made. The patient was treated with very low doses of cyclosporine A with no evidence of disease recurrence over the following 8 years. However, the discontinuation of cyclosporine A determined a relapse of the disease. The relevance of this case is due to the rarity of the disease in Italy, to its peculiar clinical presentation and, moreover, it is the first case in literature that has a good response to treatment with low doses of cyclosporine A, documented in an 8-year follow-up.

     

Comparison between VDR analogs and current immunosuppressive drugs in relation to CXCL10 secretion by human renal tubular cells

During kidney allograft rejection, CXC chemokine ligand 10 (CXCL10)–CXC chemokine receptor 3 (CXCR3) trafficking between peripheral blood and tissues initiates alloresponse and perpetuates a self‐inflammatory loop; thus, CXCL10–CXCR3 axis could represent a pharmacologic target. In this perspective, immunosuppressors targeting graft‐resident cells, beside immune cells, could be very advantageous. Vitamin D receptor (VDR) agonists exhibit considerable immunomodulatory properties. This study aimed to investigate whether elocalcitol and BXL‐01‐0029 could decrease the expression of CXCL10 in activated renal tubular cells in vitro and thus be useful in kidney allograft rejection treatment. Experiments were performed in human tubular renal cells stimulated with interferon‐γ + tumor necrosis factor‐α with and without VDR agonists, tacrolimus, sirolimus, hydrocortisone, methylprednisolone, cyclosporin A and mycophenolate mofetil. CXCL10 protein secretion and gene expression were measured by ELISA and by quantitative PCR. Specific inhibitors were used to investigate intracellular pathways involved in tubular cells activation. For IC₅₀ determination and comparison, dose‐response curves with VDR agonists, tacrolimus and mycophenolic acid were performed. Elocalcitol and BXL‐01‐0029 inhibited CXCL10 secretion by renal cells, without affecting cell viability, while almost all the immunosuppressors were found to be ineffective, except for tacrolimus and mycophenolate mofetil. BXL‐01‐0029 was the most potent drug and, notably, it was found to be capable of allowing reduction in tacrolimus‐inhibitory doses. Our data suggest that BXL‐01‐0029 could potentially be a dose‐reducing agent for conventional immunosuppressors in kidney rejection management.