Single-run reversed-phase HPLC method for determining sertraline content,enantiomeric purity,and related substances in drug substance and finished product

A direct enantio-, diastereo-, and chemo-selective high-performance liquid chromatographic method was developed for determining the content, enantiomeric purity, and related substances of the chiral antidepressant drug sertraline HCl in a single chromatographic run. The separation was achieved on a chiral stationary phase based on amylose tris(3-chloro-5-methylphenylcarbamate) under reversed-phase conditions. The method was optimized by evaluating the influence of the temperature and mobile phase composition on the retention and selectivity. The application of the single-run approach allowed to baseline resolve all investigated species in less than 15 min, without using buffers or tandem-coupled columns. The chromatographic method was validated according to the guidelines of the Official Medicines Control Laboratory and applied to control the content of sertraline HCl and related chiral substances in a generic antidepressant formulation.     

Insecure Attachment Attitudes in the Onset of Problematic Internet Use Among Late Adolescents

Studies on the role played by attachment attitudes among late adolescents who show Problematic Internet Use (PIU) are still lacking. Three self-report measures concerning attachment attitudes, childhood experiences of abuse, and Internet addiction were administered to 310 students (49 % males) aged 18–19 attending the last year of high school. Students who screened positive for PIU were more likely to be male and to have suffered childhood experiences of physical and sexual abuse; they also scored higher than the other participants on scales assessing anxious and avoidant attachment attitudes. A logistic regression showed that the classification of participants in the PIU group was predicted by male gender, having suffered from physical and sexual abuse in childhood, and preoccupation with relationships. Keeping constant the effects of gender and childhood experiences of abuse in the equation model, increasing values of preoccupation with relationships were reflected by an exponential growth in the probability curve for PIU classification. Findings of the study support the hypothesis that insecure attachment attitudes (particularly the preoccupation with relationships) are involved in the development of PIU among late adolescents.     

Brain conditioning is instrumental for successful microglia reconstitution following hematopoietic stem cell transplantation

The recent hypothesis that postnatal microglia are maintained independently of circulating monocytes by local precursors that colonize the brain before birth has relevant implications for the treatment of various neurological diseases, including lysosomal storage disorders (LSDs), for which hematopoietic cell transplantation (HCT) is applied to repopulate the recipient myeloid compartment, including microglia, with cells expressing the defective functional hydrolase. By studying wild-type and LSD mice at diverse time-points after HCT, we showed the occurrence of a short-term wave of brain infiltration by a fraction of the transplanted hematopoietic progenitors, independently from the administration of a preparatory regimen and from the presence of a disease state in the brain. However, only the use of a conditioning regimen capable of ablating functionally defined brain-resident myeloid precursors allowed turnover of microglia with the donor, mediated by local proliferation of early immigrants rather than entrance of mature cells from the circulation.     

What have we learned about antiphospholipid syndrome from patients and antiphospholipid carrier cohorts?

Venous or arterial thrombosis or pregnancy morbidity in the presence of circulating antiphospholipid antibodies (aPL) define the antiphospholipid syndrome (APS). In terms of accepted APS criteria, aPL are detected by one coagulation test (lupus anticoagulant; LAC) and two immunoassays (anticardiolipin antibodies and anti-β2-glycoptrotein I antibodies). In patients with APS, a single positive test carries a much lower risk of thrombosis recurrence or new pregnancy loss than does multiple (or triple) positivity. The same holds true for aPL carriers, namely subjects with laboratory tests but without clinical criteria for APS. Thus, very different risk categories exist among patients with APS as well as in carriers of aPL. Triple positivity apparently identifies the pathogenic autoantibody (antidomain I-II of β2-glycoptrotein I); it is in this category of patients that trials on new therapeutic strategies should focus.     

Cystic echinococcosis in sheep in Sardinia. Changing pattern and present status

The epidemiological status of cystic echinococcosis (CE) in sheep in Sardinia over the 20 years since the last control attempt at the end of the 1980s has been assessed, comparing the results of two surveys carried out in abattoirs in southern Sardinia. In the first, conducted in 1995-1997 (5-7 years after the last control effort), CE prevalence of about 75% was observed in the 1375 sheep sampled, with intensity of 10.0 and mean abundance of 7.5. The most affected organ was the liver, whereas a large percentage of infected animals presented cysts in both the liver and lung. Overall, about 26% of parasitized sheep were found to be heavily infected and 12.15% of infected animals harboured fertile cysts. In the second survey, carried out ten years after the first, during the period 2005-2010 in absence of specific control measures, a total of 1414 sheep were examined. CE prevalence was 65%, 78% in the most rural Oristano province and 58% in the most "urbanised" province of Cagliari. Frequency of sheep infected in both the liver and lung had decreased slightly compared to the first survey, particularly in the Cagliari province, as had intensity and mean abundance, though to a lesser extent in the Oristano province. 14.6% of parasitized sheep were heavily infected, showing a general decline over the 10 years between the two surveys, particularly in the Cagliari area where the figure had more than halved. Conversely, about 14% of infected sheep hosted at least 1 fertile cyst, a slight increase compared to the 1st survey. Finally, the trend of CE transmission in Sardinian sheep according to surveys carried out from 1952 to 2010 has been analysed and the results are discussed in the light of the significant socio-economic and structural transformations that have actually modified the zoonosis scenario on the island.     

Simple genetic diagnosis of hairy cell leukemia by sensitive detection of the BRAF-V600E mutation

Hairy cell leukemia (HCL) is a distinct clinicopathologic entity that responds well to purine analogs but is sometimes difficult to differentiate from HCL-like disorders (e.g., splenic marginal zone lymphoma and HCL variant). We recently identified the BRAF-V600E mutation as the disease-defining genetic event in HCL. In this study, we describe a new, simple, and inexpensive test for genetics-based diagnosis of HCL in whole-blood samples that detects BRAF-V600E through a sensitive allele-specific PCR qualitative assay followed by agarose-gel electrophoresis. This approach detected BRAF-V600E in all 123 leukemic HCL samples investigated containing as few as 0.1% leukemic cells. BRAF-V600E was detected at different time points during the disease course, even after therapy, pointing to its pivotal role in HCL pathogenesis and maintenance of the leukemic clone. Conversely, 115 non-HCL chronic B-cell neoplasms, including 79 HCL-like disorders, were invariably negative for BRAF-V600E. This molecular assay is a powerful tool for improving the diagnostic accuracy in HCL.     

Body mass index correlates with waist circumference in school aged Italian children

This study demonstrates the existence of a linear correlation between Body Mass Index (BMI) and waist circumference in Italian school aged children and suggests an indirect method (from weight and height) to estimate waist circumference, whose increase may be indicative for the diagnosis of the metabolic syndrome.     

Cyclodextrin Protects Podocytes in Diabetic Kidney Disease

Diabetic kidney disease (DKD) remains the most common cause of end-stage kidney disease despite multifactorial intervention. We demonstrated that increased cholesterol in association with downregulation of ATP-binding cassette transporter ABCA1 occurs in normal human podocytes exposed to the sera of patients with type 1 diabetes and albuminuria (DKD⁺) when compared with diabetic patients with normoalbuminuria (DKD⁻) and similar duration of diabetes and lipid profile. Glomerular downregulation of ABCA1 was confirmed in biopsies from patients with early DKD (n = 70) when compared with normal living donors (n = 32). Induction of cholesterol efflux with cyclodextrin (CD) but not inhibition of cholesterol synthesis with simvastatin prevented podocyte injury observed in vitro after exposure to patient sera. Subcutaneous administration of CD to diabetic BTBR (black and tan, brachiuric) ob/ob mice was safe and reduced albuminuria, mesangial expansion, kidney weight, and cortical cholesterol content. This was followed by an improvement of fasting insulin, blood glucose, body weight, and glucose tolerance in vivo and improved glucose-stimulated insulin release in human islets in vitro. Our data suggest that impaired reverse cholesterol transport characterizes clinical and experimental DKD and negatively influences podocyte function. Treatment with CD is safe and effective in preserving podocyte function in vitro and in vivo and may improve the metabolic control of diabetes.Diabetic kidney disease (DKD) is responsible for nearly half of the incidents of end-stage kidney disease in the U.S. (1), yet our current understanding of the pathophysiological processes responsible for DKD has led to limited improvements in patient outcomes. Multifactorial intervention reduces the rate of progression of DKD but does not prevent end-stage kidney disease in type 1 (T1D) or type 2 diabetes (T2D) (2,3). A key factor for this translation gap is the current lack of adequate mechanistic insight into DKD in humans.The kidney glomerulus is a highly specialized structure that ensures the selective ultrafiltration of plasma so that essential proteins are retained in the blood (4). Podocytes are glomerular epithelial cells that contribute to the glomerular filtration barrier through a tight regulation of actin cytoskeleton remodeling (4). Currently, the diagnosis of DKD relies on the detection of microalbuminuria (5). However, a growing body of evidence suggests that key histological lesions precede the development of albuminuria (6,7); among them, decreased podocyte number (podocytopenia) has been described as an independent predictor of DKD progression (8–12). Although we have previously shown that podocyte insulin resistance and susceptibility to apoptosis is already present at the time of onset of microalbuminuria in experimental models of DKD, the cause of podocyte injury in early DKD remains unknown (13).We used a previously established cell-based assay in which differentiated human podocytes are exposed to 4% patient sera for 24 h (14) to identify new pathways and targets in DKD. Podocytes exposed to the sera of patients with DKD showed increased cholesterol accumulation in association with downregulation of ATP-binding cassette transporter 1 (ABCA1) expression that was independent of circulating cholesterol.ABCA1 is a major regulator of cellular cholesterol homeostasis by mediating efflux to lipid-poor apolipoprotein acceptors in the bloodstream (15). ABCA1 genetic variants are strongly associated with the risk of coronary artery disease (16). Furthermore, the capacity of patient sera to induce ABCA1-mediated cholesterol efflux in macrophages is impaired in patients with T2D and incipient or overt nephropathy (17). Excessive cholesterol accumulation has been described in glomeruli of rodent models of T1D and T2D (18–20) and may contribute to DKD development and progression. Finally, inactivating mutations of ABCA1 result in Tangier disease, which causes premature atherosclerosis and proteinuria (21).Although interventions that increase ABCA1 expression (such as liver X receptor agonists) may be beneficial in DKD, they have a relatively high incidence of adverse events (22) as well as intrinsic lipogenic effects (23). We used β-cyclodextrins, cyclic oligosaccharides consisting of seven β(1-4)-glucopyranose rings, to remove cholesterol from differentiated human podocytes in vitro and from diabetic animals in vivo. The exact mechanism by which cyclodextrins (CDs) remove cholesterol from cells is not completely understood, but the formation of cholesterol/CD inclusion complexes at the membrane surface plays an important role in this process (24).We hypothesized that 2-hydroxypropyl-β-cyclodextrin, which was recently approved by the U.S. Food and Drug Administration (FDA) for the cure of Niemann-Pick disorder (25,26), would be an effective way to sequester cholesterol and to protect podocytes from cholesterol-dependent damage in DKD in vivo and in vitro.