A novel method for DNA delivery into bacteria using cationic copolymers

Amphiphilic copolymers have a wide variety of medical and biotechnological applications, including DNA transfection in eukaryotic cells. Still, no polymer-primed transfection of prokaryotic cells has been described. The reversible addition-fragmentation chain transfer (RAFT) polymer synthesis technique and the reversible deactivation radical polymerization variants allow the design of polymers with well-controlled molar mass, morphology, and hydrophilicity/hydrophobicity ratios. RAFT was used to synthesize two amphiphilic copolymers containing different ratios of the amphiphilic poly[2-(dimethyl-amino) ethyl methacrylate] and the hydrophobic poly [methyl methacrylate]. These copolymers bound to pUC-19 DNA and successfully transfected non-competent Escherichia coli DH5α, with transformation efficiency in the range of 10³ colony-forming units per µg of plasmid DNA. These results demonstrate prokaryote transformation using polymers with controlled amphiphilic/hydrophobic ratios.     

Reducing unnecessary cross-matching: a patient-specific blood ordering system is more accurate in predicting who will receive a blood transfusion than the maximum blood ordering system

Most blood transfusions are given in the operating room. Adoption of the Maximum Surgical Blood Ordering Schedule in the 1970s reduced the amount of blood unnecessarily cross-matched, but the national cross-match-to-transfusion ratio remains at approximately two-to-one. We tested the ability of a patient-specific blood ordering system (PSBOS) to more accurately predict potential operative transfusion. All adult patients who had blood cross-matched before surgery (February through June 1999) for elective operative procedures at the University of Michigan Hospital were identified. Complex surgeries were excluded. Surgeons estimated the expected blood loss for their surgeries, and the expected postoperative hematocrit was calculated using the patient's blood volume, the surgeon-defined expected blood loss, and preoperative hematocrit. Lowest tolerated hematocrit was set at 21% except in patients with coronary artery disease or who were ASA physical status III or more (28%). Sensitivity, specificity, positive predictive value, and negative predictive value of the PSBOS were calculated. Our analysis included 178 cases in which blood was cross-matched before surgery, representing 69 different surgeries and 42 surgeons. Only 16% of patients received an intraoperative transfusion. Of the 156 patients that PSBOS predicted would not require an operating room transfusion, 139 were not transfused. Of the 21 patients PSBOS predicted would be transfused, 11 were. The sensitivity of the algorithm as tested was 41%, the specificity 93%, the positive predictive value was 55%, and the negative predictive value was 89%. We conclude that PSBOS, which includes patient and surgeon variables in transfusion prediction, is more accurate than the Maximum Surgical Blood Ordering Schedule, which uses only surgical procedure. IMPLICATIONS: Currently, many units of blood set aside for surgery are never required, resulting in extra work and expense for blood banks. A formula that included patient weight and hematocrit and typical surgery blood loss was used to predict who would require transfusions. We reduced the predicted number of patients who had blood set aside from 178 to 21.     

Validity of cuff-uroflow as a diagnostic technique for bladder outlet obstruction in males

OBJECTIVE: To investigate the validity of the cuff-uroflow method as a diagnostic technique for bladder outlet obstruction (BOO) in males. MATERIAL AND METHODS: A cross-sectional study was carried out on a consecutive series of 93 males (mean age 54.1 years) referred on presenting with lower urinary tract symptoms. The patients were subjected to measurement of the isovolumetric pressure (Pcuff.op) and its corresponding flow (Qcuff.op) by means of the cuff-uroflow method. Subsequently, a study was carried out of the conventional pressure-flow, calculating the degree of BOO according to the provisional International Continence Society (ICS) nomogram. RESULTS: There was a statistical relationship between the Pcuff.op and Qcuff.op values and the degree of BOO according to the provisional ICS nomogram: patients with BOO had a lower value of Qcuff.op and a higher value of Pcuff.op than those without BOO. Using a logistic regression model a nomogram was constructed to link Pcuff.op with Qcuff.op. This nomogram was divided into three areas (obstructed, indeterminate and not obstructed). Most patients (44.1% of the sample) were situated in the indeterminate area. If those patients were excluded, the nomogram gave a sensitivity for the diagnosis of obstruction of 100% and a specificity of 55.6%. The overall rate of correct diagnosis was 84.6%. CONCLUSIONS: The parameters obtained using the cuff-uroflow method were related to the presence of BOO. According to the provisional ICS nomogram, this method showed a better sensitivity than specificity for the diagnosis of BOO.     

The central fibroblast growth factor receptor/beta klotho system: Comprehensive mapping in Mus musculus and comparisons to nonhuman primate and human samples using an automated in situ hybridization platform

Central activation of fibroblast growth factor (FGF) receptors regulates peripheral glucose homeostasis and reduces food intake in preclinical models of obesity and diabetes. The current work was undertaken to advance our understanding of the receptor expression, as sites of ligand action by FGF19, FGF21, and FGF1 in the mammalian brain remains unresolved. Recent advances in automated RNAscope in situ hybridization and droplet digital PCR (ddPCR) technology allowed us to interrogate central FGFR/beta klotho (Klb) system at the cellular level in the mouse, with relevant comparisons to nonhuman primate and human brain. FGFR1-3 gene expression was broadly distributed throughout the CNS in Mus musculus, with FGFR1 exhibiting the greatest heterogeneity. FGFR4 expression localized only in the medial habenula and subcommissural organ of mice. Likewise, Klb mRNA was restricted to the suprachiasmatic nucleus (SCh) and select midbrain and hindbrain nuclei. ddPCR in the rodent hypothalamus confirmed that, although expression levels are indeed low for Klb, there is nonetheless a bonafide subpopulation of Klb+ cells in the hypothalamus. In NHP and human midbrain and hindbrain, Klb + cells are quite rare, as is expression of FGFR4. Collectively, these data provide the most robust central map of the FGFR/Klb system to date and highlight central regions that may be of critical importance to assess central ligand effects with pharmacological dosing, such as the putative interactions between the endocrine FGFs and FGFR1/Klb, or FGF19 with FGFR4.     

Effect of chronic antipsychotic treatment on striatal phosphodiesterase 10A levels: a [11C]MP-10 PET rodent imaging study with ex vivo confirmation

A number of phosphodiesterase 10A (PDE10) inhibitors are about to undergo clinical evaluation for their efficacy in treating schizophrenia. As phosphodiesterases are in the same signalling pathway as dopamine D₂ receptors, it is possible that prior antipsychotic treatment could influence these enzyme systems in patients. Chronic, in contrast to acute, antipsychotic treatment has been reported to increase brain PDE10A levels in rodents. The aim of this study was to confirm these findings in a manner that can be translated to human imaging studies to understand its consequences. Positron emission tomography (PET) scanning was used to evaluate PDE10A enzyme availability, after chronic haloperidol administration, using a specific PDE10A ligand ([¹¹C]MP-10). The binding of [¹¹C]MP-10 in the striatum and the cerebellum was measured in rodents and a simplified reference tissue model (SRTM) with cerebellum as the reference region was used to determine the binding potential (BP_ND). In rats treated chronically with haloperidol (2 mg kg⁻¹ per day), there was no significant difference in PDE10A levels compared with the vehicle-treated group (BP_ND±s.d.: 3.57±0.64 versus 2.86±0.71). Following PET scans, ex vivo analysis of striatal brain tissue for PDE10A mRNA (Pde10a) and PDE10A enzyme activity showed no significant difference. Similarly, the PDE10A protein content determined by western blot analysis was similar between the two groups, contrary to an earlier finding. The results of the study indicate that prior exposure to antipsychotic medication in rodents does not alter PDE10A levels.     

Female genital mutilation and infections: a systematic review of the clinical evidence

Aim

Female genital mutilation (FGM) is a common practice especially performed in women with no anaesthesia or antibiotics and in absence of aseptic conditions. The aim of this systematic review is to explore and analyze for first time in the current literature, the clinical evidence related to the presence of infections in the practice of FGM.

Method

A systematic search of PubMed and Scopus was performed. A combination of the terms “female circumcision”, “genital mutilation”, “genital cutting” and “infection” were used. Studies reporting data on the infections related to patients with FGM were included.

Results

A total of 22,052 patients included, in the study, from African countries. The age ranged from 10 days to 20 years. The procedure was done by physicians, paramedical staff, and other specialties. Type I FGM was performed in 3,115 women while 5,894, 4,049 and 93 women underwent Type II, Type III and unknown type of FGM, respectively. Different types of infections were identified including UTIs, genitourinary tract infections, abscess formation and septicemia or even HIV infection. Moreover, most infections were identified in Type III FGM. The isolated pathogens in the different type of infections, were HIV, Clostridium tetani, Chlamydia trachomatis, Neisseria gonorrhoeae, Treponema pallidum, Candida albicans, Trichomonas vaginalis, HSV-2, Pseudomonas pyocyanea, Staphylococcus aureus. The univariate risk of infection ranged from 0.47 to 5.2.

Conclusion

A variety of infections can occur after FGM. The management of these complications in a low-income economy can be a great burden for the families.     

Impact of sickle cell disease and thalassemias in infants on birth outcomes

Objective

The contribution of sickle cell disease (SCD) and other common thalassemias in infants to adverse birth outcomes is under-studied. We therefore sought to compare adverse birth outcomes in infants with and without hemoglobinopathy.

Study design

Retrospective cohort study utilizing a population-based dataset from Florida (1998–2007, n = 1,564,038). The primary outcomes were low birthweight (LBW), very low birthweight (VLBW), preterm birth (PTB), very preterm birth (VPTB) and small for gestational age (SGA). We used propensity scores to match infants with hemoglobinopathy to those without hemoglobinopathy on selected variables. To approximate relative risks, we generated adjusted odds ratios (AOR) and 95% confidence intervals (CI) from logistic regression models and accounted for the matched design using generalized estimating equations framework.

Results

Infants with SCD or thalassemia had a heightened risk for LBW (AOR = 1.58, 95% CI: 1.29–1.93), VLBW (AOR = 3.01, 95% CI: 2.12–4.25), PTB (AOR = 1.36, 95% CI: 1.12–1.65), VPTB (AOR = 2.70, 95% CI: 1.93–3.78), and neurological conditions (AOR = 2.04, 95% CI: 1.48–2.81) compared to infants without hemoglobinopathy.

Conclusion

Infants with SCD or thalassemia experience considerably higher risks for multiple infant morbidities. Our findings are potentially important in prenatal counseling, as well as for targeted care of affected pregnancies in the prenatal period.     

Are prediction equations for glomerular filtration rate useful for the long-term monitoring of type 2 diabetic patients?

Background. The aim of this study was to compare the accuracyof prediction equations [modification of diet in renal disease(MDRD), simplified MDRD, Cockcroft–Gault (CG), reciprocalof creatinine and creatinine clearance] in a cohort of patientswith type 2 diabetes. Methods. A total of 525 glomerular filtration rates (GFRs) using¹²⁵I-iothalamate were carried out over 10 years in 87 type 2diabetic patients. Accuracy was evaluated at three levels ofrenal function according to the baseline values obtained withthe isotopic method: hyperfiltration (GFR: >140 ml/min/1.73m²; 140 isotopic determinations in 27 patients), normal renalfunction (GFR: 140–90 ml/min/1.73 m²; 294 isotopic determinationsin 47 patients) and chronic kidney disease (CKD) stages 2–3(GFR: 30–89 ml/min/1.73 m²; 87 isotopic determinationsin 13 patients). The annual slope for GFR (change in GFR expressedas ml/min/year) was considered to ascertain the variabilityin the equations compared with the isotopic method during follow-up.Student's t-test was used to determine the existence of significantdifferences between prediction equations and the isotopic method(P < 0.05 with Bonferroni adjusted for five contrast tests). Results. In the subgroup of patients with hyperfiltration, aGFR slope calculated with ¹²⁵I-iothalamate –4.8 ±4.7 ml/min/year was obtained. GFR slope in patients with normalrenal function was –3.0 ± 2.3 ml/min/year. In bothsituations, all equations presented a significant underestimationcompared with the isotopic GFR (P < 0.01; P < 0.05). Inthe subgroup of CKD stages 2–3, the slope for GFR with¹²⁵I-iothalamate was –1.4 ± 1.8 ml/min/year. Thebest prediction equation compared with the isotopic method provedto be MDRD with a slope for GFR of –1.4 ± 1.3 ml/min/year(P: NS) compared with the CG formula –1.0 ± 0.9ml/min/year (P: NS). Creatinine clearance presented the greatestvariability in estimation (P < 0.001). Conclusions. In the normal renal function and hyperfiltrationgroups, none of the prediction equations demonstrated acceptableaccuracy owing to excessive underestimation of renal function.In CKD stages 2–3, with mean serum creatinine 133 µmol/l(1.5 mg/dl), the MDRD equation can be used to estimate GFR duringthe monitoring and follow-up of patients with type 2 diabetesreceiving insulin, anti-diabetic drugs or both.     

Factores pronósticos del tratamiento endoscópico del reflujo vesicoureteral en pacientes con lesión medular

IntroductionVesicoureteral reflux (VUR) is an important complication in patients with spinal cord injury due to its frequency and morbidity. One of the most extended therapeutic options is endoscopic injection of obliteration substances in the urethral meatus.ObjectiveTo analyze the prognostic factors of VUR treatment using obliterative substances in patients with spinal cord injury.Material and methodsA prospective study was performed in a cohort of 76 patients (age 48.9 ± 14.4 years), of both genders, with spinal cord injuries, who underwent endoscopic treatment of the VUR during the years 2008 to 2011. In all the patients, a clinical history was obtained and a pre-operative videourodynamic study was performed. Another study was carried out at 7.32 months (standard deviation: 6.28 months) of the intervention. Treatment consisted in endoscopic injection of dextranomer/hyaluronic acid copolymer (62 cases) and polydimethylsiloxane (14). The statistical tests applied were the Fisher's exact test and the Student's T test comparing the means. Bilateral significance level was established at 95%.ResultsResolution of VUR was achieved in 46 cases (61%). The statistically significant prognostic factors were age (younger aging cured patients), bilaterality and reflects great (greater grade in bilaterality in the cases with persistence of reflux) and presence of neurogenic detrusor overactivity (greater percentage in the cases of reflux persistence). Stress urinary incontinence (greater percentage in cured patients), contractile potency (lower percentage in cured patients) and urethral resistance (greater percentage in cured patients) were also statistically significant prognostic factors.ConclusionsAmong the prognostic factors that affected the endoscopic treatment results of the VUR in patients with neurogenic lower urinary tract dysfunction (NLUTD), anatomical as well as functional factors were found.