Rapid DNA Extraction Protocol from Stool, Suitable for Molecular Genetic Diagnosis of Colon Cancer

ABSTRACT Background: Colorectal cancer (CRC) is one of the most common forms of cancers in the world and is curable if diagnosed at the early stage. Analysis of DNA extracted from stool specimens is a recent advantage to cancer diagnostics. Many protocols have been recommended for DNA extraction from stool, and almost all of them are difficult and time consuming, dealing with high amount of toxic materials like phenol. Their results vary due to sample collection method and further purification treatment. In this study, an easy and rapid method was optimized for isolating the human DNA with reduced PCR inhibitors present in stool. Methods: Fecal samples were collected from 10 colonoscopy-negative adult volunteers and 10 patients with CRC. Stool (1 g) was extracted using phenol/chloroform based protocol. The amplification of P53 exon 9 was examined to evaluate the extraction efficiency for human genomic targets and also compared its efficiency with Machiels et al. and Ito et al. protocols. Results: The amplification of exon 9 of P53 from isolated fecal DNA was possible in most cases in 35 rounds of PCR using no additional purification procedure for elimination of the remaining inhibitors. Conclusion: A useful, rapid and easy protocol for routine extraction of DNA from stool was introduced and compared with two previous protocols. Iran. Biomed. J. 1

Can Discord Domain-Containing Receptor 2 Mutation Act as a Disease Modifier for PRKAR1A Associated Melanotic Schwannoma?

Melanotic Schwannomas are rare neural sheath tumors with distinctive findings of both Schwann cells and melanocytic cells. Recognition of this entity has prompted the importance of distinction from similar tumor types such as melanomas. Early diagnosis facilitates removal of the mass with less risk of local invasion and metastasis. Although previously known as mostly benign lesions, malignant conversion and recurrence are recognized. This paper presents a patient with melanotic schwannoma, describes the distinctive features that will separate it from melanoma, and addresses the possibility of further guided therapy through next-generation sequencing

Cerebral vein thrombosis in women using short course oral contraceptive consumption

Background: Muslim women tend to postpone their menstrual period using short course oral contraceptives (OC) during Ramadan, Muslims fasting month. Recently, many cases of stroke, particularly cerebral vein thrombosis (CVT), have been reported in Ramadan. Objective: This work studies the incidence of CVT and its relation to Ramadan. Materials and Methods: This is a cohort study conducted from October 2006 to October 2009 and included 4 consecutive Ramadan’s. All patients with definite diagnosis of CVT who were referred to the neurology department of Ghaem Hospital, Mashhad, Iran in this four-year period were included in the study. Results: During the study period, 70 cases with CVT (11 males and 59 females) were recruited. Twenty five cases were admitted during Ramadan months which was significantly higher than all the other 32 months (p=1.9×10-7). The higher frequency of females (6 times more) among CVT cases probed by investigating possible risk factors and short term OC consumption was revealed as the major risk factor (p=0.00071). Conclusion: Higher incidence of CVT in females during Ramadan suggests the presence of specific risk factors in this group. Our study revealed that short-term use of OCs may be a major risk factor for CVT

Saposin C Protects Glucocerebrosidase against α‑Synuclein Inhibition

Mutations in <i>GBA1</i>, the gene for glucocerebrosidase (GCase), are genetic risk factors for Parkinson disease (PD). α-Synuclein (α-Syn), a protein implicated in PD, interacts with GCase and efficiently inhibits enzyme activity. GCase deficiency causes the lysosomal storage disorder Gaucher disease (GD). We show that saposin C (Sap C), a protein vital for GCase activity <i>in vivo</i>, protects GCase against α-syn inhibition. Using nuclear magnetic resonance spectroscopy, site-specific fluorescence, and Förster energy transfer probes, Sap C was observed to displace α-syn from GCase in solution and on lipid vesicles. Our results suggest that Sap C might play a crucial role in GD-related PD

Identification of miRNAs that modulate glucocerebrosidase activity in Gaucher disease cells

<div><p>Gaucher disease is an autosomal recessive disorder caused by deficiency of the enzyme glucocerebrosidase. Although it is a monogenic disease, there is vast phenotypic heterogeneity, even among patients with the same genotype. MicroRNAs (miRNAs) are small non-coding RNAs involved in many biological processes and diseases. To determine whether miRNAs can affect glucocerebrosidase activity, we performed a screen of 875 different miRNA mimics. The screen was performed using Gaucher fibroblasts, and glucocerebrosidase activity was used as the initial outcome parameter. We found several miRNAs that either up- or down-regulated glucocerebrosidase activity. In follow-up assays, we confirmed that one specific miRNA (miR-127–5p) down-regulated both glucocerebrosidase activity and protein levels by down-regulation of LIMP-2, the receptor involved in proper trafficking of glucocerebrosidase from the endoplasmic reticulum to the lysosome. A conditioned media assay demonstrated that cells treated with this miRNA secreted glucocerebrosidase into the extracellular environment, supporting impaired LIMP-2 function. Two other miRNAs, miR-16–5p and miR-195–5p, were found to up-regulate glucocerebrosidase activity by greater than 40% and to enhance expression and protein levels of the enzyme. In conclusion, we show that miRNAs can alter glucocerebrosidase activity in patient cells, indicating that miRNAs can potentially act as modifiers in Gaucher disease.</p></div