Archive for December, 2009

Mutation screen and association studies for the fatty acid amide hydrolase (FAAH) gene and early onset and adult obesity

Thursday, December 31st, 2009

Background:
The orexigenic effects of cannabinoids are limited by activation of the endocannabinoid degrading enzyme fatty acid amide hydrolase (FAAH). The aim of this study was to analyse whether FAAH alleles are associated with early and late onset obesity.
Methods:
We initially assessed association of five single nucleotide polymorphisms (SNPs) in FAAH with early onset extreme obesity in up to 521 German obese children and both parents. SNPs with nominal p-values [less than or equal to] 0.1 were subsequently analysed in 235 independent German obesity families. SNPs associated with childhood obesity (p-values[less than or equal to]0.05) were further analysed in 8,491 adult individuals of a population-based cohort (KORA) for association with adult obesity. One SNP was further analysed in 985 German obese adults and 588 normal and underweight controls. In parallel, we screened the FAAH coding region for novel sequence variants in 92 extremely obese children using single-stranded-conformation-polymorphism-analysis and denaturing HPLC and assessed the implication of the identified new variants for childhood obesity.
Results:
The trio analysis revealed some evidence for an association of three SNPs in FAAH (rs324420, rs324419 and rs873978) with childhood obesity (two-sided p-values between 0.06 and 0.10). Although analyses of these variants in 235 independent obesity families did not result in statistically significant effects (two-sided p-values between 0.14 and 0.75), the combined analysis of all 603 obesity families supported the idea of an association of two SNPs in FAAH (rs324420 and rs2295632) with early onset extreme obesity (p-values between 0.02 and 0.03). No association was, however, found between these variants and adult obesity. The mutation screen revealed four novel variants, which were not associated with early onset obesity (p>0.05).
Conclusions:
As we observed some evidence for an association of the FAAH variants rs2295632, rs324420 with early onset but not adult obesity, we conclude that the FAAH variants analyzed here at least do not seem to play a major role in the etiology of obesity within our samples.

Effects of chronic low dose rotenone treatment on human microglial cells

Wednesday, December 30th, 2009

Background:
Exposure to toxins / chemicals is considered to be a significant risk factor in the pathogenesis of Parkinson’s disease (PD); one putative chemical is the naturally occurring herbicide rotenone that is now used widely in establishing PD models. We, and others, have shown that chronic low dose rotenone treatment induces excessive accumulation of Reactive Oxygen Species (ROS), inclusion body formation and apoptosis in dopaminergic neurons of animal and human origin. Some studies have also suggested that microglia enhance the rotenone induced neurotoxicity. While the effects of rotenone on neurons are well established, there is little or no information available on the effect of rotenone on microglial cells, and especially cells of human origin. The aim of the present study was to investigate the effects of chronic low dose rotenone treatment on human microglial CHME-5 cells.
Methods:
We have shown previously that rotenone induced inclusion body formation in human dopaminergic SH-SY5Y cells and therefore used these cells as a control for inclusion body formation in this study. SH-SY5Y and CHME-5 cells were treated with 5nM rotenone for four weeks. At the end of week 4, both cell types were analysed for the presence of inclusion bodies, superoxide dismutases and cell activation (only in CHME-5 cells) using Haematoxylin and Eosin staining, immunocytochemical and western blotting methods. Levels of active caspases and ROS (both extra and intra cellular) were measured using biochemical methods.
Conclusion:
The results suggest that chronic low dose rotenone treatment activates human microglia (cell line) in a manner similar to microglia of animal origin as shown by others. However human microglia release excessive amounts of ROS extracellularly, do not show excessive amounts of intracellular ROS and active caspases and most importantly do not show any protein aggregation or inclusion body formation. Human microglia appear to be resistant to rotenone (chronic, low dose) induced damage.

Multilocus variable-number tandem repeat analysis for molecular typing and phylogenetic analysis of Shigella flexneri

Wednesday, December 30th, 2009

Background:
Shigella flexneri is one of the causative agents of shigellosis, a major cause of childhood mortality in developing countries. Multilocus variable-number tandem repeat (VNTR) analysis (MLVA) is a prominent subtyping method to resolve closely related bacterial isolates for investigation of disease outbreaks and provide information for establishing phylogenetic patterns among isolates. The present study aimed to develop an MLVA method for S. flexneri and the VNTR loci identified were tested on 242 S. flexneri isolates to evaluate their variability in various serotypes. The isolates were also analyzed by pulsed-field gel electrophoresis (PFGE) to compare the discriminatory power and to investigate the usefulness of MLVA as a tool for phylogenetic analysis of S. flexneri.
Results:
Thirty-six VNTR loci were identified by exploring the repeat sequence loci in genomic sequences of Shigella species and by testing the loci on nine isolates of different subserotypes. The VNTR loci in different serotype groups differed greatly in their variability. The discriminatory power of an MLVA assay based on four most variable VNTR loci was higher, though not significantly, than PFGE for the total isolates, a panel of 2a isolates, which were relatively diverse, and a panel of 4a/Y isolates, which were closely-related. Phylogenetic groupings based on PFGE patterns and MLVA profiles were considerably concordant. The genetic relationships among the isolates were correlated with serotypes. The phylogenetic trees constructed using PFGE patterns and MLVA profiles presented two distinct clusters for the isolates of serotype 3 and one distinct cluster for each of the serotype groups, 1a/1b/NT, 2a/2b/X/NT, 4a/Y, and 6. Isolates that had different serotype but had closer genetic relatedness than those with the same serotype were observed between serotype Y and subserotype 4a, serotype X and subserotype 2b, subserotype 1a and 1b, and subserotype 3a and 3b.
Conclusions:
The 36 VNTR loci identified exhibited considerably different degrees of variability among S. flexneri serotype groups. VNTR locus could be highly variable in a serotype but invariable in others. MLVA assay based on four highly variable loci could display a comparable resolving power to PFGE in discriminating isolates. MLVA is also a prominent molecular tool for phylogenetic analysis of S. flexneri; the resulting data are beneficial to establish clear clonal patterns among different serotype groups and to discern clonal groups among isolates within the same serotype. As highly variable VNTR loci could be serotype-specific, a common MLVA protocol that consists of only a few loci, for example four to eight loci, and that provides high resolving power to all S. flenxeri serotypes may not be obtainable.

Chronic fungal sinusitis leading to disastrous cerebral aspergillosis: a case report

Wednesday, December 30th, 2009

Cerebral angioinvasion is a fatal complication of disseminated aspergillosis and has been rarely described in diabetic population in the absence of ketoacidosis. We present a case of invasive fungal sinusitis in a 79 year old diabetic man who presented with chronic frontal headaches. Despite appropriate medical and surgical treatment, the disease progressed and the patient eventually succumbed to a fatal ruptured mycotic aneurysm. We emphasize that clinicians should consider this in the differential diagnoses of all diabetics who present with chronic sinusitis, as early diagnosis could be the key in the successful treatment.

Torsion of ovarian cyst during pregnancy: a case report

Wednesday, December 30th, 2009

In this case we report a 23 -year-old primigravida with 30 weeks presenting with torsion of the ovarian cyst. She presented to the antenatal clinic with acute pain abdomen. She was diagnosed to have torsion of ovarian cyst during pregnancy and a cystecomy was carried out. Her histopathology report showed a benign serous cystadenoma. Her pregnancy was followed up. She delivered a healthy female baby at term. Although the safety of antepartum surgical intervention has been accepted, abdominal surgery nevertheless carries some risks to a pregnant woman and unborn fetus, and so the choice of management necessitates a weighing of risks based on characterization of the adnexal mass and gestational age.