Cloning, expression and characterization of l-asparaginase from Withania somnifera L. for large scale production

l-Asparaginase (E.C. 3.5.1.1) is used as a therapeutic agent in the treatment of acute childhood lymphoblastic leukemia. It is found in a variety of organisms such as microbes, plants and mammals. In plants, l-asparaginase enzymes are required to catalyze the release of ammonia from asparagine, which is the main nitrogen-relocation molecule in these organisms. An Indian medicinal plant, Withania somnifera was reported as a novel source of l-asparaginase. l-Asparaginase from W. somnifera was cloned and overexpressed in E. coli. The enzymatic properties of the recombinant enzyme were investigated and the kinetic parameters (Km, kcat) for a number of substrates were determined. The kinetic parameters of selected substrates were determined at various pH and the pH- and temperature-dependence profiles were analyzed. WA gene successfully cloned into E. coli BL21 (DE3) showed high asparaginase activity with a specific activity of 17.3 IU/mg protein

Physicians' attitudes towards ePrescribing – evaluation of a Swedish full-scale implementation

<p>Abstract</p> <p>Background</p> <p>The penetration rate of Electronic Health Record (EHR) systems in health care is increasing. However, many different EHR-systems are used with varying ePrescription designs and functionalities. The aim of the present study was to evaluate experienced ePrescribers' attitudes towards ePrescribing for suggesting improvements.</p> <p>Methods</p> <p>Physicians (n = 431) from seven out of the 21 Swedish health care regions, using one of the six most widely implemented EHR-systems with integrated electronic prescribing modules, were recruited from primary care centers and hospital clinics of internal medicine, orthopaedics and surgery. The physicians received a web survey that comprised eight questions on background data and 19 items covering attitudes towards ePrescribing. Forty-two percent (n = 199) of the physicians answered the questionnaire; 90% (n = 180) of the respondents met the inclusion criteria and were included in the final analysis.</p> <p>Results</p> <p>A majority of the respondents regarded their EHR-system easy to use in general (81%), and for the prescribing of drugs (88%). Most respondents believed they were able to provide the patients better service by ePrescribing (92%), and regarded ePrescriptions to be time saving (91%) and to be safer (83%), compared to handwritten prescriptions. Some of the most frequently reported weaknesses were: not clearly displayed price of drugs (43%), complicated drug choice (21%), and the perception that it was possible to handle more than one patient at a time when ePrescribing (13%). Moreover, 62% reported a lack of receipt from the pharmacy after successful transmission of an ePrescription. Although a majority (73%) of the physicians reported that they were always or often checking the ePrescription a last time before transmitting, 25% declared that they were seldom or never doing a last check. The respondents suggested a number of improvements, among others, to simplify the drug choice and the cancellation of ePrescriptions.</p> <p>Conclusion</p> <p>The Swedish physicians in the group studied were generally satisfied with their specific EHR-system and with ePrescribing as such. However, identified weaknesses warrant improvements of the EHR-systems as well as of their implementation in the individual health care organisation.</p

Urinary exosome miR-146a is a potential marker of albuminuria in essential hypertension

BACKGROUND: There is increasing interest in using extracellular vesicle-derived microRNAs (miRNAs) as biomarkers in renal dysfunction and injury. Preliminary evidence indicates that miRNAs regulate the progression of glomerular disease. Indeed, exosomes from the renal system have provided novel evidence in the clinical setting of albuminuria. Thus, the aim of this study was to quantify the urinary miRNAs present in exosome and microvesicles (MVs), and to assess their association with the presence of increased urinary albumin excretion in essential hypertension. METHODS: Exosomes were collected from urine specimens from a cohort of hypertensive patients with (n = 24) or without albuminuria (n = 28), and from 20 healthy volunteers as a control group. Urinary exosomes were phenotyped by Western blot, tunable resistive pulse sensing, and electronic microscopy. Expression of miR-146a and miR-335* was analysed by qRT-PCR and any associations between albuminuria and exosomal miRNAs were analysed. RESULTS: Urinary miRNAs are highly enriched in exosome subpopulations compared to MVs, both in patients with or without increased albuminuria (p < 0.001), but not in the control group. High albuminuria was associated with 2.5-fold less miR-146a in exosomes (p = 0.017), whereas miR-146a levels in MV did not change. In addition, exosome miR-146a levels were inversely associated with albuminuria (r = 0.65, p < 0.0001), and discriminated the presence of urinary albumin excretion presence [area under the curve = 0.80, 95% confidence interval: 0.66-0.95; p = 0.0013]. CONCLUSIONS: Our results indicate that miRNAs were enriched in the urinary exosome subpopulation in hypertensive patients and that low miR-146a expression in exosomes was associated with the presence of albuminuria. Thus, urinary exosome miR-146a may be a potentially useful tool for studying early renal injury in hypertension

Circulating microRNAs as novel biomarkers for diabetes mellitus.

Diabetes mellitus is characterized by insulin secretion from pancreatic β cells that is insufficient to maintain blood glucose homeostasis. Autoimmune destruction of β cells results in type 1 diabetes mellitus, whereas conditions that reduce insulin sensitivity and negatively affect β-cell activities result in type 2 diabetes mellitus. Without proper management, patients with diabetes mellitus develop serious complications that reduce their quality of life and life expectancy. Biomarkers for early detection of the disease and identification of individuals at risk of developing complications would greatly improve the care of these patients. Small non-coding RNAs called microRNAs (miRNAs) control gene expression and participate in many physiopathological processes. Hundreds of miRNAs are actively or passively released in the circulation and can be used to evaluate health status and disease progression. Both type 1 diabetes mellitus and type 2 diabetes mellitus are associated with distinct modifications in the profile of miRNAs in the blood, which are sometimes detectable several years before the disease manifests. Moreover, circulating levels of certain miRNAs seem to be predictive of long-term complications. Technical and scientific obstacles still exist that need to be overcome, but circulating miRNAs might soon become part of the diagnostic arsenal to identify individuals at risk of developing diabetes mellitus and its devastating complications